Antibody-drug conjugate T-DM1 treatment for HER2+ breast cancer induces ROR1 and confers resistance through activation of Hippo transcriptional coactivator YAP1

Abstract

Background: A newly developed drug trastuzumab emtansine (T-DM1) has improved the survival of breast cancer (BC) patients. Despite an impressive initial clinical response, a subgroup of patient develop resistance and present therapeutic challenges. The underlying resistance mechanisms are not fully investigated. We report that T-DM1 treatment modulates the expression of ROR1 (type 1 receptor tyrosine kinase-like orphan receptor)and induces self-renewal of cancer stem cells (CSCs) leading to therapeutic resistance.

Methods: Using BC patient tumor samples, and BC cell lines we gained insight into the T-DM1 treatment inducedROR1 over expression and resistance.In vitrosphere forming assays and in vivo extreme dilution assays were employed to analyze the stemness and self-renewal capacity of the cells. A series of molecular expression and protein assays including qRT-PCR, FACS-sorting, ELISA, immunostaining, Western blotting were used to provide evidence.

Findings: Exposure of cells to T-DM1 shifted ROR1 expression from low to high, enriched within the CSC subpopulation, coincident with increased Bmi1 and stemness factors. T-DM1 induced ROR1 cells showed high spheroidand tumor forming efficiency in vitro and in an animal model exhibiting shorter tumor-free time.Mechanistically,the over expression of ROR1 is partly induced by the activation of YAP1 and its target genes. Silencing of ROR1 andYAP1 by pharmacologic inhibitors and/or sh/siRNA inhibited spheroid formation, the initiation of tumors and the capacity for self-renewal and ROR1 over expression.

Interpretations: The results presented here indicate that simultaneous targeting of ROR1 and YAP1 may suppress CSC self-renewal efficacy and inhibit tumor progression in BC. In this manner such treatments may overcome theT-DM1 mediated therapeutic resistance and improve clinical outcome.