RESUMEN
BACKGROUND: Triple negative breast cancers (TNBC) have poor prognoses despite aggressive treatment with cytotoxic chemotherapy. Cancer-associated fibroblasts (CAFs) are prominent in tumour stroma. Our hypothesis was that CAFs modulate chemotherapy sensitivity. METHODS: TNBC cells and breast fibroblasts were cultured; survival after chemotherapeutics was assessed using luciferase or clonogenic assays. Signalling was investigated using transcriptomics, reporters, recombinant proteins and blocking antibodies. Clinical relevance was investigated using immunohistochemistry. RESULTS: Breast CAFs dose-dependently protected TNBC cell lines MDA-MB-231 and MDA-MB-157, but not MDA-MB-468s, from chemotherapy. CAF-induced protection was associated with interferon (IFN) activation. CAFs were induced to express IFNß1 by chemotherapy and TNBC co-culture, leading to paracrine activation in cancer cells. Recombinant IFNs were sufficient to protect MDA-MB-231 and MDA-MB-157 but not MDA-MB-468 cells. In TNBC patients, IFNß1 expression in CAFs correlated with cancer cell expression of MX1, a marker of activated IFN signalling. High expression of IFNß1 (CAFs) or MX1 (tumour cells) correlated with reduced survival after chemotherapy, especially in claudin-low tumours (which MDA-MB-231 and MDA-MB-157 cells represent). Antibodies that block IFN receptors reduced CAF-dependent chemoprotection. CONCLUSIONS: CAF-induced activation of IFN signalling in claudin-low TNBCs results in chemoresistance. Inhibition of this pathway represents a novel method to improve breast cancer outcomes.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Fibroblastos Asociados al Cáncer/patología , Fibroblastos/patología , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Interferón beta/metabolismo , Proteínas de Resistencia a Mixovirus/metabolismo , Neoplasias de la Mama Triple Negativas/patología , Apoptosis , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Fibroblastos Asociados al Cáncer/efectos de los fármacos , Fibroblastos Asociados al Cáncer/metabolismo , Proliferación Celular , Técnicas de Cocultivo , Femenino , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Humanos , Interferón beta/genética , Proteínas de Resistencia a Mixovirus/genética , Comunicación Paracrina , Pronóstico , Transcriptoma , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Neoplasias de la Mama Triple Negativas/genética , Neoplasias de la Mama Triple Negativas/metabolismo , Células Tumorales CultivadasRESUMEN
MicroRNAs and RNA-binding proteins exert regulation on >60% of coding genes, yet interplay between them is little studied. Canonical microRNA binding occurs by base-pairing of microRNA 3'-ends to complementary "seed regions" in mRNA 3'UTRs, resulting in translational repression. Similarly, regulatory RNA-binding proteins bind to mRNAs, modifying stability or translation. We investigated post-transcriptional regulation acting on the xenobiotic pump ABCB1/P-glycoprotein, which is implicated in cancer therapy resistance. We characterised the ABCB1 UTRs in primary breast cancer cells and identified UTR sequences that responded to miR-19b despite lacking a canonical binding site. Sequences did, however, contain consensus sites for the RNA-binding protein HuR. We demonstrated that a tripartite complex of HuR, miR-19b and UTR directs repression of ABCB1/P-glycoprotein expression, with HuR essential for non-canonical miR-19b binding thereby controlling chemosensitivity of breast cancer cells. This exemplifies a new cooperative model between RNA-binding proteins and microRNAs to expand the repertoire of mRNAs that can be regulated. This study suggests a novel therapeutic target to impair P-glycoprotein mediated drug efflux, and also indicates that current microRNA binding predictions that rely on seed regions alone may be too conservative.