The Interfollicular Epidermis of Adult Mouse Tail Comprises Two Distinct Cell Lineages that Are Differentially Regulated by Wnt, Edaradd, and Lrig1

Current models of how mouse tail interfollicular epidermis (IFE) is maintained overlook the coexistence of two distinct terminal differentiation programs: parakeratotic (scale) and orthokeratotic (interscale). Lineage tracing and clonal analysis revealed that scale and interscale are maintained by unipotent cells in the underlying basal layer, with scale progenitors dividing more rapidly than interscale progenitors. Although scales are pigmented and precisely aligned with hair follicles, melanocytes and follicles were not necessary for scale differentiation. Epidermal Wnt signaling was required for scale enlargement during development and for postnatal maintenance of scale-interscale boundaries. Loss of Edaradd inhibited ventral scale formation, whereas loss of Lrig1 led to scale enlargement and fusion. In wild-type skin, Lrig1 was not expressed in IFE but was selectively upregulated in dermal fibroblasts underlying the interscale. We conclude that the different IFE differentiation compartments are maintained by distinct stem cell populations and are regulated by epidermal and dermal signals

An immune response gene expression module identifies a good prognosis subtype in estrogen receptor negative breast cancer

A feature selection method was used in an analysis of three major microarray expression datasets to identify molecular subclasses and prognostic markers in estrogen receptor-negative breast cancer, showing that it is a heterogeneous disease with at least four main subtypes

Genomic evidence supports a clonal diaspora model for metastases of esophageal adenocarcinoma.

The poor outcomes in esophageal adenocarcinoma (EAC) prompted us to interrogate the pattern and timing of metastatic spread. Whole-genome sequencing and phylogenetic analysis of 388 samples across 18 individuals with EAC showed, in 90% of patients, that multiple subclones from the primary tumor spread very rapidly from the primary site to form multiple metastases, including lymph nodes and distant tissues-a mode of dissemination that we term 'clonal diaspora'. Metastatic subclones at autopsy were present in tissue and blood samples from earlier time points. These findings have implications for our understanding and clinical evaluation of EAC.MRC core grant (RG84369), an NIHR Research Professorship (RG67258) and Cancer Research UK (RG66287)

Transcriptomic profiling reveals three molecular phenotypes of adenocarcinoma at the gastroesophageal junction.

Cancers occurring at the gastroesophageal junction (GEJ) are classified as predominantly esophageal or gastric, which is often difficult to decipher. We hypothesized that the transcriptomic profile might reveal molecular subgroups which could help to define the tumor origin and behavior beyond anatomical location. The gene expression profiles of 107 treatment-naïve, intestinal type, gastroesophageal adenocarcinomas were assessed by the Illumina-HTv4.0 beadchip. Differential gene expression (limma), unsupervised subgroup assignment (mclust) and pathway analysis (gage) were undertaken in R statistical computing and results were related to demographic and clinical parameters. Unsupervised assignment of the gene expression profiles revealed three distinct molecular subgroups, which were not associated with anatomical location, tumor stage or grade (p > 0.05). Group 1 was enriched for pathways involved in cell turnover, Group 2 was enriched for metabolic processes and Group 3 for immune-response pathways. Patients in group 1 showed the worst overall survival (p = 0.019). Key genes for the three subtypes were confirmed by immunohistochemistry. The newly defined intrinsic subtypes were analyzed in four independent datasets of gastric and esophageal adenocarcinomas with transcriptomic data available (RNAseq data: OCCAMS cohort, n = 158; gene expression arrays: Belfast, n = 63; Singapore, n = 191; Asian Cancer Research Group, n = 300). The subgroups were represented in the independent cohorts and pooled analysis confirmed the prognostic effect of the new subtypes. In conclusion, adenocarcinomas at the GEJ comprise three distinct molecular phenotypes which do not reflect anatomical location but rather inform our understanding of the key pathways expressed.We would like to acknowledge The Human Research Tissue Bank which is supported by the NIHR Cambridge Biomedical Research Centre ... This work has been supported by the research scholarship BO4097/1‐1 from the Deutsche Forschungsgemeinschaft (DFG) for JB, grant RG67258 of the National Institute for Health and Research (NIHR) and grant RG66287 of Cancer Research UK (CRUK) have been awarded to RCF

Organoid cultures recapitulate esophageal adenocarcinoma heterogeneity providing a model for clonality studies and precision therapeutics

Esophageal adenocarcinoma (EAC) incidence is increasing while 5-year survival rates remain less than 15%. A lack of experimental models has hampered progress. We have generated clinically annotated EAC organoid cultures that recapitulate the morphology, genomic and transcriptomic landscape of the primary tumor including point mutations, copy number alterations and mutational signatures. Karyotyping has confirmed polyclonality reflecting the clonal architecture of the primary and subclones underwent clonal selection associated with driver gene status. Medium throughput drug sensitivity testing demonstrates the potential of targeting receptor tyrosine kinases and downstream mediators. EAC organoid cultures provide a pre-clinical tool for studies of clonal evolution and precision therapeutics

Mutational signatures in esophageal adenocarcinoma define etiologically distinct subgroups with therapeutic relevance.

Esophageal adenocarcinoma (EAC) has a poor outcome, and targeted therapy trials have thus far been disappointing owing to a lack of robust stratification methods. Whole-genome sequencing (WGS) analysis of 129 cases demonstrated that this is a heterogeneous cancer dominated by copy number alterations with frequent large-scale rearrangements. Co-amplification of receptor tyrosine kinases (RTKs) and/or downstream mitogenic activation is almost ubiquitous; thus tailored combination RTK inhibitor (RTKi) therapy might be required, as we demonstrate in vitro. However, mutational signatures showed three distinct molecular subtypes with potential therapeutic relevance, which we verified in an independent cohort (n = 87): (i) enrichment for BRCA signature with prevalent defects in the homologous recombination pathway; (ii) dominant T>G mutational pattern associated with a high mutational load and neoantigen burden; and (iii) C>A/T mutational pattern with evidence of an aging imprint. These subtypes could be ascertained using a clinically applicable sequencing strategy (low coverage) as a basis for therapy selection.Whole-genome sequencing of esophageal adenocarcinoma samples was performed as part of the International Cancer Genome Consortium (ICGC) through the oEsophageal Cancer Clinical and Molecular Stratification (OCCAMS) Consortium and was funded by Cancer Research UK. We thank the ICGC members for their input on verification standards as part of the benchmarking exercise. We thank the Human Research Tissue Bank, which is supported by the National Institute for Health Research (NIHR) Cambridge Biomedical Research Centre, from Addenbrooke’s Hospital and UCL. Also the University Hospital of Southampton Trust and the Southampton, Birmingham, Edinburgh and UCL Experimental Cancer Medicine Centres and the QEHB charities. This study was partly funded by a project grant from Cancer Research UK. R.C.F. is funded by an NIHR Professorship and receives core funding from the Medical Research Council and infrastructure support from the Biomedical Research Centre and the Experimental Cancer Medicine Centre. We acknowledge the support of The University of Cambridge, Cancer Research UK (C14303/A17197) and Hutchison Whampoa Limited. We would like to thank Dr. Peter Van Loo for providing the NGS version of ASCAT for copy number calling. We are grateful to all the patients who provided written consent for participation in this study and the staff at all participating centres. Some of the work was undertaken at UCLH/UCL who received a proportion of funding from the Department of Health’s NIHR Biomedical Research Centres funding scheme. The work at UCLH/UCL was also supported by the CRUK UCL Early Cancer Medicine Centre.This is the author accepted manuscript. The final version is available from Nature Publishing Group via http://dx.doi.org/10.1038/ng.365

Patient-specific cancer genes contribute to recurrently perturbed pathways and establish therapeutic vulnerabilities in esophageal adenocarcinoma

Abstract: The identification of cancer-promoting genetic alterations is challenging particularly in highly unstable and heterogeneous cancers, such as esophageal adenocarcinoma (EAC). Here we describe a machine learning algorithm to identify cancer genes in individual patients considering all types of damaging alterations simultaneously. Analysing 261 EACs from the OCCAMS Consortium, we discover helper genes that, alongside well-known drivers, promote cancer. We confirm the robustness of our approach in 107 additional EACs. Unlike recurrent alterations of known drivers, these cancer helper genes are rare or patient-specific. However, they converge towards perturbations of well-known cancer processes. Recurrence of the same process perturbations, rather than individual genes, divides EACs into six clusters differing in their molecular and clinical features. Experimentally mimicking the alterations of predicted helper genes in cancer and pre-cancer cells validates their contribution to disease progression, while reverting their alterations reveals EAC acquired dependencies that can be exploited in therapy

Immune activation by DNA damage predicts response to chemotherapy and survival in oesophageal adenocarcinoma.

OBJECTIVE: Current strategies to guide selection of neoadjuvant therapy in oesophageal adenocarcinoma (OAC) are inadequate. We assessed the ability of a DNA damage immune response (DDIR) assay to predict response following neoadjuvant chemotherapy in OAC. DESIGN: Transcriptional profiling of 273 formalin-fixed paraffin-embedded prechemotherapy endoscopic OAC biopsies was performed. All patients were treated with platinum-based neoadjuvant chemotherapy and resection between 2003 and 2014 at four centres in the Oesophageal Cancer Clinical and Molecular Stratification consortium. CD8 and programmed death ligand 1 (PD-L1) immunohistochemical staining was assessed in matched resection specimens from 126 cases. Kaplan-Meier and Cox proportional hazards regression analysis were applied according to DDIR status for recurrence-free survival (RFS) and overall survival (OS). RESULTS: A total of 66 OAC samples (24%) were DDIR positive with the remaining 207 samples (76%) being DDIR negative. DDIR assay positivity was associated with improved RFS (HR: 0.61; 95% CI 0.38 to 0.98; p=0.042) and OS (HR: 0.52; 95% CI 0.31 to 0.88; p=0.015) following multivariate analysis. DDIR-positive patients had a higher pathological response rate (p=0.033), lower nodal burden (p=0.026) and reduced circumferential margin involvement (p=0.007). No difference in OS was observed according to DDIR status in an independent surgery-alone dataset.DDIR-positive OAC tumours were also associated with the presence of CD8+ lymphocytes (intratumoural: p<0.001; stromal: p=0.026) as well as PD-L1 expression (intratumoural: p=0.047; stromal: p=0.025). CONCLUSION: The DDIR assay is strongly predictive of benefit from DNA-damaging neoadjuvant chemotherapy followed by surgical resection and is associated with a proinflammatory microenvironment in OAC.This work was supported by the Gastrointestinal Cancer Research Charitable Fund administered by the Belfast Health and Social Care Trust, the Cancer Research UK Experimental Cancer Medicine Centre Initiative, Invest Northern Ireland and Almac Diagnostics. Oesophageal Cancer Clinical and Molecular Stratification (OCCAMS) was funded by a programme grant from Cancer Research UK (RG66287). We would like to thank the Human Research Tissue Bank, which is supported by the National Institute for Health Research (NIHR) Cambridge Biomedical Research Centre from Addenbrooke’s Hospital. Additional infrastructure support was provided from the CRUK funded Experimental Cancer Medicine Centre. RF has programmatic funding from the Medical Research Council and infrastructure support from the NIHR Biomedical Research Centre and the Cambridge Experimental Medicine Centre. Tissue samples used in this research were received from the Northern Ireland Biobank, which is funded by HSC Research and Development Division of the Public Health Agency in Northern Ireland and Cancer Research UK through the Belfast Cancer Research UK Centre and the Northern Ireland Experimental Cancer Medicine Centre; additional support was received from the Friends of the Cancer Centre. The Northern Ireland Molecular Pathology Laboratory has received funding from Cancer Research UK, the Friends of the Cancer Centre and the Sean Crummey Foundation. This project has received funding from the European Union’s Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grant agreement no 721906. The OCCAMS Study Group is a multicentre UK collaboration