LAUSR

Background: 10-15 signal transduction pathways govern major cellular processes, e.g. cell division, differentiation and migration, both in physiology and pathophysiology. They are frequently abnormally active in cancer and can drive cancer growth and metastasis. Resistance to targeted drugs directed towards specific signaling pathways like the ER pathway, can be mediated by induction of activity of other signaling pathways. The past decade we developed a new method to quantitatively measure functional activity status of signal transduction pathways in individual cell and tissue samples, based on Bayesian computational model inference of pathway activity from measurements of mRNA levels of target genes of the pathway-associated transcription factor1,2,3. Assays have been developed for androgen (AR) and estrogen receptor (ER), Hedgehog (HH), Wnt, TGFb, Notch, NFkB, PI3K, JAK-STAT 1/2 and 3, and MAPK-AP1 pathways. They can be performed simultaneously on a single cell or tissue sample using Affymetrix HG-U133 Plus 2.0 microarray or qPCR, and provide quantitative pathway activity scores expressed on a log2odds scale. Method. Pathway analysis using these assays was performed on public Affymetrix expression microarray data (GSE74391,4) from ER-positive MCF-7 cells that were either sensitive or resistant to fulvestrant (n=26). Results. Resistance to fulvestrant was associated with complete loss of functional ER pathway activity (difference 16 log2odds score, p=0.00029) and gain of MAPK-AP1 pathway activity (difference 5.8 log2odds, p=0.00029) in ER positive MCF-7 cells. In the resistant MCF-7 cells, PI3K and MAPK-AP1 pathway activity were negatively correlated (Pearson 0.652, p=0.002), suggesting mutually exclusive pathway activation. Conclusion. Our pathway assays enabled identification of the previously unidentified mechanism of resistance to fulvestrant in a cell culture model system for breast cancer, demonstrating that: (1) ER positivity is a prerequisite, but not sufficient, for ER pathway activity, in agreement with our earlier clinical studies1,5,6;; (2) MAPK pathway activity may confer resistance to hormonal therapy; (3) in the absence of MAPK-AP1 pathway activity, the PI3K pathway may be an alternative resistance pathway; (3) signalling pathway assays are well suited to (quantitatively) investigate activity of signal transduction pathways which may confer resistance to a (targeted) drug treatment. References 1. Verhaegh, W. et al. SCancer Res. 74, 2936–2945 (2014). 2. van Ooijen, H. et al.Am. J. Pathol. 188, 1956–1972 (2018). 3. Stolpe, A. van de, et al. Sci Rep 9, 1603 (2019). 4. Alves, C. L. et al.Clin. Cancer Res. 22, 5514–5526 (2016). 5. Blok, E. J. et al.Cancer Res 76, P3-07-65-P3-07–65 (2016). 6. Yang, S.-R. et al.Cancer Res 79, P5-11-06-P5-11–06 (2019). Citation Format: Yvonne Wesseling-Rozendaal, Anja van de Stolpe, Laurent Holtzer, Marcia Alves de Inda, Paul van de Wiel. Fulvestrant resistance in an MCF-7 model for breast cancer is associated with complete loss of ER pathway activity and gain of MAPK-AP1 pathway activity [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr A131. doi:10.1158/1535-7163.TARG-19-A131