BackgroundRecurrence after >5-year disease-free survival affects one-fifth of breast cancer patients and is the clinical manifestation of cancer cell reactivation after persistent dormancy.MethodsWe investigated cellular dormancy in vitro and in vivo using breast cancer cell lines and cell and molecular biology techniques.ResultsWe demonstrated cellular dormancy in breast cancer bone metastasis, associated with haematopoietic stem cell (HSC) mimicry, in vivo competition for HSC engraftment and non-random distribution of dormant cells at the endosteal niche. Notch2 signal implication was demonstrated by immunophenotyping the endosteal niche-associated cancer cells and upon co-culture with sorted endosteal niche cells, which inhibited breast cancer cell proliferation in a Notch2-dependent manner. Blocking this signal by in vivo acute administration of the γ-secretase inhibitor, dibenzazepine, induced dormant cell mobilisation from the endosteal niche and colonisation of visceral organs. Sorted Notch2HIGH breast cancer cells exhibited a unique stem phenotype similar to HSCs and in vitro tumour-initiating ability in mammosphere assay. Human samples confirmed the existence of a small Notch2HIGH cell population in primary and bone metastatic breast cancers, with a survival advantage for Notch2HIGH vs Notch2LOW patients.ConclusionsNotch2 represents a key determinant of breast cancer cellular dormancy and mobilisation in the bone microenvironment.

Oesophageal adenocarcinoma (OAC) patients show poor survival rates and there are few targeted molecular therapies available. However, components of the receptor tyrosine kinase (RTK) driven pathways are commonly mutated in OAC, typified by high frequency amplifications of the RTK ERBB2. ERBB2 can be therapeutically targeted, but this has limited clinical benefit due to the acquisition of drug resistance. Here we examined how OAC cells adapt to ERBB2 inhibition as they transition to a drug resistant state. ERBB2 inhibition triggers widespread remodelling of the accessible chromatin landscape and the underlying gene regulatory networks. The transcriptional regulators HNF4A and PPARGC1A play a key role in this network rewiring. Initially, inhibition of cell cycle associated gene expression programmes is observed, with compensatory increases in the programmes driving changes in metabolic activity. Both PPARGC1A and HNF4A are required for the acquisition of resistance to ERBB2 inhibition and PPARGC1A is instrumental in promoting a switch to dependency on oxidative phosphorylation. Our work therefore reveals the molecular pathways that support the acquisition of a resistant state and points to potential new therapeutic strategies to combat cellular adaptation and ensuing drug resistance.

Oesophageal adenocarcinoma (OAC) patients show poor survival rates and there are few targeted molecular therapies available. However, components of the receptor tyrosine kinase (RTK) driven pathways are commonly mutated in OAC, typified by high frequency amplifications of the RTK ERRB2. ERBB2 can be therapeutically targeted, but this has limited clinical benefit due to the acquisition of drug resistance. Here we examined how OAC cells respond to ERBB2 inhibition through altering their regulatory chromatin landscapes and rewiring their gene regulatory networks to acquire a reversible resistant state. ERBB2 inhibition triggers widespread remodelling of the accessible chromatin landscape. This remodelling is accompanied by the activation of the transcriptional regulators HNF4A and PPARGC1A. Initially, inhibition of cell cycle associated gene expression programmes is observed, with compensatory increases in the programmes driving changes in metabolic activity. PPARGC1A is instrumental in promoting a switch to dependency on oxidative phosphorylation and both PPARGC1A and HNF4A are required for the acquisition of resistance to ERBB2 inhibition. Our work therefore reveals the molecular pathways that support the acquisition of a resistant state and points to potential new therapeutic strategies to combat drug resistance.