Background:Oestrogen receptor-negative (ER−) breast cancer is intrinsically sensitive to chemotherapy. However, tumour response is often incomplete, and relapse occurs with high frequency. The aim of this work was to analyse the molecular characteristics of residual tumours and early response to chemotherapy in patient-derived xenografts (PDXs) of breast cancer.Methods:Gene and protein expression profiles were analysed in a panel of ER− breast cancer PDXs before and after chemotherapy treatment. Tumour and stromal interferon-gamma expression was measured in xenografts lysates by human and mouse cytokine arrays, respectively.Results:The analysis of residual tumour cells in chemo-responder PDX revealed a strong overexpression of IFN-inducible genes, induced early after AC treatment and associated with increased STAT1 phosphorylation, DNA-damage and apoptosis. No increase in IFN-inducible gene expression was observed in chemo-resistant PDXs upon chemotherapy. Overexpression of IFN-related genes was associated with human IFN-γ secretion by tumour cells.Conclusions:Treatment-induced activation of the IFN/STAT1 pathway in tumour cells is associated with chemotherapy response in ER− breast cancer. Further validations in prospective clinical trials will aim to evaluate the usefulness of this signature to assist therapeutic strategies in the clinical setting.

Activation of the IFN/STAT1 pathway is closely associated with drug response and recurrence of breast cancer treated by chemotherapy. The aim of the current study was to elucidate the molecular mechanisms involved upstream and downstream of this pathway in order to identify distinct entities that might be manipulated to improve treatment efficacy. Four breast cancer cell lines (T-47D, MCF7, MDA-MB-231 and HBCx-19 established from the eponymous PDX) were treated in vitro with mafosfamide, a DNA damage inducer. In two of these cell lines (MCF7 and HBCx-19), genotoxic treatment upregulated type I IFN expression leading to paracrine activation of IFN/STAT1 signaling pathway after 6–8 days. We show that STING, a well-characterized inducer of IFN in immune cells, is rapidly triggered in MCF7 cells under genotoxic stress and forms nuclear foci that co-localize with phosphorylated IRF-3 and γH2AX. STING silencing abrogated chemotherapy-induced type I IFN production and signaling and potentiated genotoxic treatment efficacy as it promoted cell death extent and delayed cell colony regrowth. Similar results were obtained after silencing PARP12, one selected gene of the IFN/STAT1 pathway fingerprint. In summary, this study provides the first demonstration of STING activation in breast cancer cells. Our data suggest that genotoxic-induced, STING-mediated type I IFN signaling is a cell-intrinsic mechanism of breast cancer cell survival and regrowth.

Triple negative breast cancer (TNBC) is a tumor subtype characterized by the absence of overexpressed estrogen receptor-alpha (ER), progesterone receptor (PR), and HER2 receptor, encoded by ERBB2, a known proto-oncogene. This type of tumors account for approximately 15–25% of breast cancers at diagnosis, and is one of the most aggressive subtypes, with 77% of patients that live free of disease 5 years post-diagnosis. One of the most reliable predictive markers of patient outcome is the pathological complete response (pCR), which indicates that the surgical specimen removed after neoadjuvant chemotherapy contains no viable tumor cells detectable at histopathological level. For patients with pCR, the probability of surviving the disease is very high, however, pCR is observed only in about 20–30% of TNBC. On the other hand, for patients with no pCR the probablity of developing recurrent disease at 5 years is 50%.\ud \ud As pCR is strongly correlated with treatment efficacy, it is mandatory to develop methods that allow to tell as quick as possible if the treatment chosen for a given patient is efficiently working or if it should be abandoned in favor of an alternative strategy that could prove more efficacious. XenTech collection of breast cancer patient-derived xenografts (PDXs) includes 25 models of TNBC that display heterogeneous response to different chemotherapy agents. We used our models to investigate if transcriptional changes could be detected in PDXs that responded well to genotoxic agents. To do this we analyzed the gene expression profile of laser-microdissected residual tumor nodules interspersed in the murine stroma upon very efficient response to Adriamycin/Cyclophosphamide (AC). When doing so, we identified several genes of the IFN/STAT1 pathway that were over-expressed when compared to untreated tumors. This activation seems to be a transient event, as it was lost in tumors relapsing after the residual tumor nodule stage.\ud \ud The finding that residual cells from tumors strongly responding to AC treatment over-expressed IFN/STAT1 pathway-related genes prompted us to investigate whether this effect could be detected as an early event upon tumor exposure to chemotherapy. All TNBC models tested that were good responders to AC treatment displayed over-expression of IFN/STAT1 pathway-related genes as early as 3 days post-treatment, most of them reaching a plateau of intensity at day 7 post-treatment. By contrast, TNBC insensitive or low responders to AC treatment failed to show over-expression of IFN/STAT1 pathway-related genes.\ud \ud To verify if the selective over-expression of these genes in TNBC models sensitive to AC was independent of the treatment administered, Irinotecan and Capecitabine were used to treat TNBC models with heterogeneous response to these drugs. Again, we found that overexpression of IFN/STAT1 pathway-related genes was specifically identified at early stages only in TNBC models that responded well to these drugs.\ud \ud These results suggest that genes of the IFN/STAT1 pathway ...

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