Abstract PD8-02: Kinome profiling of ER+ breast cancer PDXs identifies PKMYT1 as a marker of hormone independent growth and poor outcome
Authors: Chen Anran , Kim Beom-Jun , Chan Doug W , Singh Purba , Dobrolecki Lacey E , Lei Jonathan T , Li Shunqiang , Welm Alana L , Lewis Michael T , Ellis Matthew J 
Published: February 2021

Background: Endocrine therapy resistance is common and is a leading cause of breast cancer-related death. Thus, the search for therapeutic targets based on mechanistic insights into endocrine therapy resistance continues. Kinases are important drug targets and regulators in cells, many of which are involved in tumorigenesis and the development of treatment resistance. Mass spectrometry-based kinome analysis has been impeded by the low abundance of individual kinases. Here we utilized kinase inhibitor-conjugated beads to enrich and thereby sensitively profile the kinome of estrogen receptor-positive (ER+) breast cancer patient-derived xenograft (PDX) tumors under estradiol-deprivation treatment.

Experimental design and methods: We harvested tumor samples from 20 breast PDX lines with various degrees of estradiol (E2) dependence in ovariectomized SCID-beige mice with and without E2 supplementation (n=3 per PDX line per arm). The kinases in the tumor lysates were enriched using kinase inhibitor pulldown (KIP) beads and the tightly bound kinases were quantified by mass spectrometry. To identify candidates, we selected for kinases that were highly E2-regulated in the E2-dependent PDX lines but constitutively expressed in E2-independent PDX lines. Survival analysis of candidate kinases in patients with ER+ breast cancer was performed using the METABRIC dataset.

Results: Each PDX line had a unique yet reproducible kinome. To seek kinases with consistent relationships with estrogen dependence, we sought kinases that were statistically differentially expressed in tumors between E2 supplied and deprived conditions. Noticeably, membrane-associated tyrosine-and threonine-specific cdc2-inhibitory kinase (PKMYT1), a WEE family kinase known to have estrogen response elements (EREs) in its regulatory region, was significantly decreased after E2 deprivation in E2-dependent PDXs (log2 fold change = -7.86, p&lt;0.001) but was constitutive in E2-independent PDXs. High PKMYT1 mRNA expression was associated with poor prognosis in the ER+ samples in METABRIC (hazard ratio=2.2, p&lt;0.001). In contrast, the more studied member of the WEE family, WEE1, lacks an estrogen response element (ERE) and is not E2-regulated from the kinome profiling result. WEE1 mRNA expression level is not associated with the outcome of patients with ER+ breast cancer, suggesting that PKMYT1 has evolved a specific role in the cell cycle of ER+ tumors.

Conclusion: Here, we analyzed the kinomes of 20 ER+ breast cancer PDX tumors with or without E2 by mass spectrometry. We discovered that PKMYT1 is a marker of hormone independent growth and poor outcome. Ongoing experiments that study the effects of PKMYT1 inhibition in both ER-dependent and independent circumstances will be presented.

Citation Format: Anran Chen, Beom-Jun Kim, Doug W Chan, Purba Singh, Lacey E Dobrolecki, Jonathan T Lei, Shunqiang Li, Alana L Welm, Michael T Lewis, Matthew J Ellis. Kinome profiling of ER+ breast cancer PDXs identifies PKMYT1 as a marker of hormone independent growth and poor outcome [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PD8-02.