Here we studied cell-free plasma DNA (cfDNA) collected from subjects with advanced lung cancer whose tumors had developed resistance to the epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) AZD9291. We first performed next-generation sequencing of cfDNA from seven subjects and detected an acquired EGFR C797S mutation in one; expression of this mutant EGFR construct in a cell line rendered it resistant to AZD9291. We then performed droplet digital PCR on serial cfDNA specimens collected from 15 AZD9291-treated subjects. All were positive for T790M prior to treatment, but at resistance three molecular subtypes emerged: 6 cases acquired the C797S mutation, 5 cases maintained the T790M mutation but did not acquire the C797S mutation, and 4 cases lost the T790M mutation despite detecting of the underlying EGFR activating mutation. Our findings provide insight into the diversity of mechanisms through which tumors acquire resistance to AZD9291 and highlight the need for therapies able to overcome resistance mediated by EGFR C797S.

Cell-free circulating tumour DNA (ctDNA) in plasma has been shown to be informative of the genomic alterations present in tumours and has been used to monitor tumour progression and response to treatments. However, patients with brain tumours do not present with or present with low amounts of ctDNA in plasma precluding the genomic characterization of brain cancer through plasma ctDNA. Here we show that ctDNA derived from central nervous system tumours is more abundantly present in the cerebrospinal fluid (CSF) than in plasma. Massively parallel sequencing of CSF ctDNA more comprehensively characterizes the genomic alterations of brain tumours than plasma, allowing the identification of actionable brain tumour somatic mutations. We show that CSF ctDNA levels longitudinally fluctuate in time and follow the changes in brain tumour burden providing biomarkers to monitor brain malignancies. Moreover, CSF ctDNA is shown to facilitate and complement the diagnosis of leptomeningeal carcinomatosis.

While tumor genome sequencing has become widely available in clinical and research settings, the interpretation of tumor somatic variants remains an important bottleneck. Here we present the Cancer Genome Interpreter, a versatile platform that automates the interpretation of newly sequenced cancer genomes, annotating the potential of alterations detected in tumors to act as drivers and their possible effect on treatment response. The results are organized in different levels of evidence according to current knowledge, which we envision can support a broad range of oncology use cases. The resource is publicly available at http://www.cancergenomeinterpreter.org.Electronic supplementary materialThe online version of this article (10.1186/s13073-018-0531-8) contains supplementary material, which is available to authorized users.

The Schizosaccharomyces pombe transcription factor Pap1 regulates antioxidant-gene transcription in response to H2O2. Pap1 activation occurs only at low, but not elevated, H2O2 concentrations that instead strongly trigger the mitogen-activated protein kinase Sty1 pathway. Here, we identify the peroxiredoxin Tpx1 as the upstream activator of Pap1. We show that, at low H2O2 concentrations, this oxidant scavenger can transfer a redox signal to Pap1, whereas higher concentrations of the oxidant inhibit the Tpx1-Pap1 redox relay through the temporal inactivation of Tpx1 by oxidation of its catalytic cysteine to a sulfinic acid. This cysteine modification can be reversed by the sulfiredoxin Srx1, its expression in response to high doses of H2O2 strictly depending on active Sty1. Thus, Tpx1 oxidation to the cysteine-sulfinic acid and its reversion by Srx1 constitutes a previously uncharacterized redox switch in H2O2 signaling, restricting Pap1 activation within a narrow range of H2O2 concentrations.Sty1 ͉ thiol oxidation ͉ H2O2 sensor ͉ Prx ͉ fission yeast

Antibody targeting of the immune checkpoint receptor PD1 produces therapeutic activity in a variety of solid tumors, but most patients exhibit partial or complete resistance to treatment for reasons that are unclear. In this study, we evaluated tumor specimens from 65 patients with melanoma, lung nonsquamous, squamous cell lung or head and neck cancers who were treated with the approved PD1-targeting antibodies pembrolizumab or nivolumab. Tumor RNA before anti-PD1 therapy was analyzed on the nCounter system using the PanCancer 730-Immune Panel, and we identified 23 immune-related genes or signatures linked to response and progression-free survival (PFS). In addition, we evaluated intra- and interbiopsy variability of PD1, PD-L1, CD8A, and CD4 mRNAs and their relationship with tumor-infiltrating lymphocytes (TIL) and PD-L1 IHC expression. Among the biomarkers examined, PD1 gene expression along with 12 signatures tracking CD8 and CD4 T-cell activation, natural killer cells, and IFN activation associated significantly with nonprogressive disease and PFS. These associations were independent of sample timing, drug used, or cancer type. TIL correlated moderately (∼0.50) with PD1 and CD8A mRNA levels and weakly (∼0.35) with CD4 and PD-L1. IHC expression of PD-L1 correlated strongly with PD-L1 (0.90), moderately with CD4 and CD8A, and weakly with PD1. Reproducibility of gene expression in intra- and interbiopsy specimens was very high (total SD <3%). Overall, our results support the hypothesis that identification of a preexisting and stable adaptive immune response as defined by mRNA expression pattern is reproducible and sufficient to predict clinical outcome, regardless of the type of cancer or the PD1 therapeutic antibody administered to patients. .

The genes encoding tyrosine kinase receptors EphB2 and EphB3 are beta-catenin and Tcf4 target genes in colorectal cancer (CRC) and in normal intestinal cells. In the intestinal epithelium, EphB signaling controls the positioning of cell types along the crypt-villus axis. In CRC, EphB activity suppresses tumor progression beyond the earliest stages. Here we show that EphB receptors compartmentalize the expansion of CRC cells through a mechanism dependent on E-cadherin-mediated adhesion. We demonstrate that EphB-mediated compartmentalization restricts the spreading of EphB-expressing tumor cells into ephrin-B1-positive territories in vitro and in vivo. Our results indicate that CRC cells must silence EphB expression to avoid repulsive interactions imposed by normal ephrin-B1-expressing intestinal cells at the onset of tumorigenesis.

The Wnt–β-catenin and PI3K-AKT-FOXO3a pathways have a central role in cancer. AKT phosporylates FOXO3a, relocating it from the cell nucleus to the cytoplasm, an effect that is reversed by PI3K and AKT inhibitors. Simultaneous hyperactivation of the Wnt–β-catenin pathway and inhibition of PI3K-AKT signaling promote nuclear accumulation of β-catenin and FOXO3a, respectively, promoting cell scattering and metastasis by regulating a defined set of target genes. Indeed, the anti-tumoral AKT inhibitor API-2 promotes nuclear FOXO3a accumulation and metastasis of cells with high nuclear β-catenin content. Nuclear β-catenin confers resistance to the FOXO3a-mediated apoptosis induced by PI3K and AKT inhibitors in patient-derived primary cultures and in corresponding xenograft tumors in mice. This resistance is reversed by XAV-939, an inhibitor of Wnt–β-catenin signaling. In the presence of high nuclear β-catenin content, activation of FOXO3a by PI3K or AKT inhibitors makes it behave as a metastasis inductor rather than a proapoptotic tumor suppressor. We show that it is possible to evaluate the β-catenin status of patients' carcinomas and the response of patient-derived cells to target-directed drugs that accumulate FOXO3a in the nucleus before deciding on a course of treatment. We propose that this evaluation could be essential to the provision of a safer and more effective personalized treatment.

Background BRCA1 and BRCA2 (BRCA1/2)-deficient tumors display impaired homologous recombination repair (HRR) and enhanced sensitivity to DNA damaging agents or to poly(ADP-ribose) polymerase (PARP) inhibitors (PARPi). Their efficacy in germline BRCA1/2 (gBRCA1/2)-mutated metastatic breast cancers has been recently confirmed in clinical trials. Numerous mechanisms of PARPi resistance have been described, whose clinical relevance in gBRCA-mutated breast cancer is unknown. This highlights the need to identify functional biomarkers to better predict PARPi sensitivity.Patients and methodsWe investigated the in vivo mechanisms of PARPi resistance in gBRCA1 patient-derived tumor xenografts (PDXs) exhibiting differential response to PARPi. Analysis included exome sequencing and immunostaining of DNA damage response proteins to functionally evaluate HRR. Findings were validated in a retrospective sample set from gBRCA1/2-cancer patients treated with PARPi.ResultsRAD51 nuclear foci, a surrogate marker of HRR functionality, were the only common feature in PDX and patient samples with primary or acquired PARPi resistance. Consistently, low RAD51 was associated with objective response to PARPi. Evaluation of the RAD51 biomarker in untreated tumors was feasible due to endogenous DNA damage. In PARPi-resistant gBRCA1 PDXs, genetic analysis found no in-frame secondary mutations, but BRCA1 hypomorphic proteins in 60% of the models, TP53BP1-loss in 20% and RAD51-amplification in one sample, none mutually exclusive. Conversely, one of three PARPi-resistant gBRCA2 tumors displayed BRCA2 restoration by exome sequencing. In PDXs, PARPi resistance could be reverted upon combination of a PARPi with an ataxia-telangiectasia mutated (ATM) inhibitor.ConclusionDetection of RAD51 foci in gBRCA tumors correlates with PARPi resistance regardless of the underlying mechanism restoring HRR function. This is a promising biomarker to be used in the clinic to better select patients for PARPi therapy. Our study also supports the clinical development of PARPi combinations such as those with ATM inhibitors.