Epigenetic homogeneity within colorectal tumors predicts shorter relapse-free and overall survival times for patients with locoregional cancer

Background & aims: there are few validated biomarkers that can be used to predict outcomes for patients with colorectal cancer. Part of the challenge is the genetic and molecular heterogeneity of colorectal tumors not only among patients, but also within tumors. We have explored intratumor heterogeneity at the epigenetic level, due to its dynamic nature. We analyzed DNA methylation profiles of the digestive tract surface and the central bulk and invasive front regions of colorectal tumors. Methods: we determined the DNA methylation profiles of >450,000 CpG sites in 3 macrodissected regions of 79 colorectal tumors and 23 associated liver metastases, obtained from 2 hospitals in Spain. We also analyzed samples for KRAS and BRAF mutations, 499,170 single nucleotide polymorphisms, and performed immunohistochemical analyses. Results: we observed differences in DNA methylation among the 3 tumor sections; regions of tumor−host interface differed the most from the other tumor sections. Interestingly, tumor samples collected from areas closer to the gastrointestinal transit most frequently shared methylation events with metastases. When we calculated individual coefficients to quantify heterogeneity, we found that epigenetic homogeneity was significantly associated with short time of relapse-free survival (log-rank P = .037) and short time of overall survival (log-rank P = .026) in patients with locoregional colorectal cancer. Conclusions: in an analysis of 79 colorectal tumors, we found significant heterogeneity in patterns of DNA methylation within each tumor; the level of heterogeneity correlates with times of relapse-free and overall survival

Single human oocyte transcriptome analysis reveals distinct maturation stage-dependent pathways impacted by age

Female fertility is inversely correlated with maternal age due to a depletion of the oocyte pool and a reduction in oocyte developmental competence. Few studies have addressed the effect of maternal age on the human mature oocyte (MII) transcriptome, which is established during oocyte growth and maturation, however, the pathways involved remain unclear. Here, we characterize and compare the transcriptomes of a large cohort of fully grown germinal vesicle stage (GV) and in vitro matured (IVM-MII) oocytes from women of varying reproductive age. First, we identified two clusters of cells reflecting the oocyte maturation stage (GV and IVM-MII) with 4445 and 324 putative marker genes, respectively. Furthermore, we identified genes for which transcript representation either progressively increased or decreased with age. Our results indicate that the transcriptome is more affected by age in IVM-MII oocytes (1219 genes) than in GV oocytes (596 genes). In particular, we found that transcripts of genes involved in chromosome segregation and RNA splicing significantly increased representation with age, while genes related to mitochondrial activity showed a lower representation. Gene regulatory network analysis facilitated the identification of potential upstream master regulators of the genes involved in those biological functions. Our analysis suggests that advanced maternal age does not globally affect the oocyte transcriptome at GV or IVM-MII stages. Nonetheless, hundreds of genes displayed altered transcript representation, particularly in IVM-MII oocytes, which might contribute to the age-related quality decline in human oocytes.Work on this study in the laboratory of B.P. has been funded by the AXA research fund (AXA Chair in Risk prediction in age-related diseases), contributions from Clinica EUGIN (Identification of Epigenetic Effects of Ageing on Human Oocytes), the Spanish Ministry of Science, Innovation and Universities (BFU2017-88407-P), the Agencia Estatal de Investigación (AEI) (EUR2019-103817) and the Catalan Agència de Gestió d'Ajuts Universitaris i de Recerca (AGAUR, 2017 SGR 346). S.L. has received funding from the European Union's Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie grant agreement No 754422. We acknowledge support of the Spanish Ministry of Science and Innovation through the Instituto de Salud Carlos III, to the EMBL partnership and to the Co-financing with funds from the European Regional Development Fund (FEDER) (Programa Operativo FEDER Plurirregional de España (POPE) 2014-2020). We also acknowledge support of the Centro de Excelencia Severo Ochoa and the Generalitat de Catalunya through the CERCA Programme, the Departament de Salut and Departament d'Empresa i Coneixement and through the Secretaria d'Universitats i Recerca for the Co-financing with FEDER funds (Programa Operatiu FEDER de Catalunya 2014-2020). Furthermore, this study has been supported by intramural funding of Clinica EUGIN to R.V

Epigenetic homogeneity within colorectal tumors predicts shorter relapse-free and overall survival times for patients with locoregional cancer

Background & aims: there are few validated biomarkers that can be used to predict outcomes for patients with colorectal cancer. Part of the challenge is the genetic and molecular heterogeneity of colorectal tumors not only among patients, but also within tumors. We have explored intratumor heterogeneity at the epigenetic level, due to its dynamic nature. We analyzed DNA methylation profiles of the digestive tract surface and the central bulk and invasive front regions of colorectal tumors. Methods: we determined the DNA methylation profiles of >450,000 CpG sites in 3 macrodissected regions of 79 colorectal tumors and 23 associated liver metastases, obtained from 2 hospitals in Spain. We also analyzed samples for KRAS and BRAF mutations, 499,170 single nucleotide polymorphisms, and performed immunohistochemical analyses. Results: we observed differences in DNA methylation among the 3 tumor sections; regions of tumor−host interface differed the most from the other tumor sections. Interestingly, tumor samples collected from areas closer to the gastrointestinal transit most frequently shared methylation events with metastases. When we calculated individual coefficients to quantify heterogeneity, we found that epigenetic homogeneity was significantly associated with short time of relapse-free survival (log-rank P = .037) and short time of overall survival (log-rank P = .026) in patients with locoregional colorectal cancer. Conclusions: in an analysis of 79 colorectal tumors, we found significant heterogeneity in patterns of DNA methylation within each tumor; the level of heterogeneity correlates with times of relapse-free and overall survival

Controlled X-chromosome dynamics defines meiotic potential of female mouse in vitro germ cells

The mammalian germline is characterized by extensive epigenetic reprogramming during its development into functional eggs and sperm. Specifically, the epigenome requires resetting before parental marks can be established and transmitted to the next generation. In the female germline, X-chromosome inactivation and reactivation are among the most prominent epigenetic reprogramming events, yet very little is known about their kinetics and biological function. Here, we investigate X-inactivation and reactivation dynamics using a tailor-made in vitro system of primordial germ cell-like cell (PGCLC) differentiation from mouse embryonic stem cells. We find that X-inactivation in PGCLCs in vitro and in germ cell-competent epiblast cells in vivo is moderate compared to somatic cells, and frequently characterized by escaping genes. X-inactivation is followed by step-wise X-reactivation, which is mostly completed during meiotic prophase I. Furthermore, we find that PGCLCs which fail to undergo X-inactivation or reactivate too rapidly display impaired meiotic potential. Thus, our data reveal fine-tuned X-chromosome remodelling as a critical feature of female germ cell development towards meiosis and oogenesis.This work was supported by the Spanish Ministry of Science, Innovation and Universities (BFU2014-55275-P, BFU2017-88407-P to B.P.), the Agencia Estatal de Investigación (AEI) (EUR2019-103817 to B.P.), the AXA Research Fund (to B.P.) and the Agencia de Gestio d’Ajuts Universitaris i de Recerca (AGAUR, 2017 SGR 346 to B.P.). We would like to thank the Spanish Ministry of Economy, Industry and Competitiveness (MEIC) to the EMBL partnership and to the “Centro de Excelencia Severo Ochoa.” We also acknowledge support of the CERCA Programme of the Generalitat de Catalunya. N.A. is supported by an EMBO postdoctoral fellowship (LTF 695-2019). J.S. and M.B. were supported by La Caixa International PhD Fellowships and J.S. by a travel grant from the Company of Biologists (Development Journal). P.A. has received funding from the European Union’s Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie grant agreement No 75165

A single-cell tumor immune atlas for precision oncology

The tumor immune microenvironment is a main contributor to cancer progression and a promising therapeutic target for oncology. However, immune microenvironments vary profoundly between patients, and biomarkers for prognosis and treatment response lack precision. A comprehensive compendium of tumor immune cells is required to pinpoint predictive cellular states and their spatial localization. We generated a single-cell tumor immune atlas, jointly analyzing published data sets of \u3e500,000 cells from 217 patients and 13 cancer types, providing the basis for a patient stratification based on immune cell compositions. Projecting immune cells from external tumors onto the atlas facilitated an automated cell annotation system. To enable in situ mapping of immune populations for digital pathology, we applied SPOTlight, combining single-cell and spatial transcriptomics data and identifying colocalization patterns of immune, stromal, and cancer cells in tumor sections. We expect the tumor immune cell atlas, together with our versatile toolbox for precision oncology, to advance currently applied stratification approaches for prognosis and immunotherapy

A single-cell tumor immune atlas for precision oncology

The tumor immune microenvironment is a main contributor to cancer progression and a promising therapeutic target for oncology. However, immune microenvironments vary profoundly between patients, and biomarkers for prognosis and treatment response lack precision. A comprehensive compendium of tumor immune cells is required to pinpoint predictive cellular states and their spatial localization. We generated a single-cell tumor immune atlas, jointly analyzing published data sets of >500,000 cells from 217 patients and 13 cancer types, providing the basis for a patient stratification based on immune cell compositions. Projecting immune cells from external tumors onto the atlas facilitated an automated cell annotation system. To enable in situ mapping of immune populations for digital pathology, we applied SPOTlight, combining single-cell and spatial transcriptomics data and identifying colocalization patterns of immune, stromal, and cancer cells in tumor sections. We expect the tumor immune cell atlas, together with our versatile toolbox for precision oncology, to advance currently applied stratification approaches for prognosis and immunotherapy.This publication is part of a project (BCLLATLAS) that has received funding from the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation programme (grant agreement No. 810287). This work has received funding from the Ministerio de Ciencia, Innovación y Universidades (SAF2017-89109-P; AEI/FEDER, UE) and the Fundació La Marató de TV3 (201903-30-31-32).We further acknowledge funding from the St. Vincent’s Clinic Foundation (V.T.C.) and the National Health and Medical Research Council Investigator Grant (APP1175781, J.E.P.), the Fundación Asociación Española contra el Cáncer (AECC), FERO (EDM), Ramón Areces Foundation, Cellex Foundation, BBVA (CAIMI), the ISCIII, FIS (PI16/01278), Juan de la Cierva formación fellowship (C.R.-P.) and Sara Borrell fellowship (E.P.-R.). Core funding is from the ISCIII and the Generalitat de Catalunya. We acknowledge support of the Spanish Ministry of Economy, Industry and Competitiveness (MEIC) to the EMBL partnership, the Centro de Excelencia Severo Ochoa, the CERCA Programme / Generalitat de Catalunya, the Spanish Ministry of Economy, Industry and Competitiveness (MEIC) through the nstituto de Salud Carlos III and the Generalitat de Catalunya through Departament de Salut and Departament d’Empresa i Coneixement. We also acknowledge the cofinancing by the Spanish Ministry of Economy, Industry and Competitiveness (MEIC) with funds from the European Regional Development Fund (ERDF) corresponding to the 2014–2020 Smart Growth Operating Progra

Mutations in JMJD1C are involved in Rett syndrome and intellectual disability

PURPOSE: Autism spectrum disorders are associated with defects in social response and communication that often occur in the context of intellectual disability. Rett syndrome is one example in which epilepsy, motor impairment, and motor disturbance may co-occur. Mutations in histone demethylases are known to occur in several of these syndromes. Herein, we aimed to identify whether mutations in the candidate histone demethylase JMJD1C (jumonji domain containing 1C) are implicated in these disorders. METHODS: We performed the mutational and functional analysis of JMJD1C in 215 cases of autism spectrum disorders, intellectual disability, and Rett syndrome without a known genetic defect. RESULTS: We found seven JMJD1C variants that were not present in any control sample ( 6,000) and caused an amino acid change involving a different functional group. From these, two de novo JMJD1C germline mutations were identified in a case of Rett syndrome and in a patient with intellectual disability. The functional study of the JMJD1C mutant Rett syndrome patient demonstrated that the altered protein had abnormal subcellular localization, diminished activity to demethylate the DNA damage-response protein MDC1, and reduced binding to MECP2. We confirmed that JMJD1C protein is widely expressed in brain regions and that its depletion compromises dendritic activity. CONCLUSIONS: Our findings indicate that mutations in JMJD1C contribute to the development of Rett syndrome and intellectual disability.Genet Med 18 1, 378-385.This study was supported by the European Community’s Seventh Framework Program (FP7/2007–2013) under grant agreement PITN-GA-2012–316758 of the EPITRAIN project and PITN-GA-2009–238242 of DISCHROM; ERC grant agreement 268626 of the EPINORC project; the E-RARE EuroRETT network (Carlos III Health Institute project PI071327); the Fondation Lejeune (France); MINECO projects SAF2011-22803 and CSD2006-00049; the Cellex Foundation; the Botín Foundation; the Catalan Association for Rett Syndrome; Fundación Alicia Koplowitz 2011 Grant AKOPLOWITZ11_006; the FIS project PI1002512; Grants PI10/01422, PI13/00285, CA10/01474, RD06/0020/1050, RD12/0036/008, and 2009-SGR293; and the Health and Science Departments of the Catalan government (Generalitat de Catalunya). K.S. and P.P. are EPITRAIN Research Fellows. M.E. is an ICREA Research Professor

Metastatic recurrence in colorectal cancer arises from residual EMP1+ cells

Around 30-40% of patients with colorectal cancer (CRC) undergoing curative resection of the primary tumour will develop metastases in the subsequent years1. Therapies to prevent disease relapse remain an unmet medical need. Here we uncover the identity and features of the residual tumour cells responsible for CRC relapse. An analysis of single-cell transcriptomes of samples from patients with CRC revealed that the majority of genes associated with a poor prognosis are expressed by a unique tumour cell population that we named high-relapse cells (HRCs). We established a human-like mouse model of microsatellite-stable CRC that undergoes metastatic relapse after surgical resection of the primary tumour. Residual HRCs occult in mouse livers after primary CRC surgery gave rise to multiple cell types over time, including LGR5+ stem-like tumour cells2-4, and caused overt metastatic disease. Using Emp1 (encoding epithelial membrane protein 1) as a marker gene for HRCs, we tracked and selectively eliminated this cell population. Genetic ablation of EMP1high cells prevented metastatic recurrence and mice remained disease-free after surgery. We also found that HRC-rich micrometastases were infiltrated with T cells, yet became progressively immune-excluded during outgrowth. Treatment with neoadjuvant immunotherapy eliminated residual metastatic cells and prevented mice from relapsing after surgery. Together, our findings reveal the cell-state dynamics of residual disease in CRC and anticipate that therapies targeting HRCs may help to avoid metastatic relapse.© 2022. The Author(s), under exclusive licence to Springer Nature Limited

Mutations in JMJD1C are involved in Rett syndrome and intellectual disability

Purpose: autism spectrum disorders are associated with defects in social response and communication that often occur in the context of intellectual disability. Rett syndrome is one example in which epilepsy, motor impairment, and motor disturbance may co-occur. Mutations in histone demethylases are known to occur in several of these syndromes. Herein, we aimed to identify whether mutations in the candidate histone demethylase JMJD1C (jumonji domain containing 1C) are implicated in these disorders. Methods: we performed the mutational and functional analysis of JMJD1C in 215 cases of autism spectrum disorders, intellectual disability, and Rett syndrome without a known genetic defect. Results: we found seven JMJD1C variants that were not present in any control sample (~ 6,000) and caused an amino acid change involving a different functional group. From these, two de novo JMJD1C germline mutations were identified in a case of Rett syndrome and in a patient with intellectual disability. The functional study of the JMJD1C mutant Rett syndrome patient demonstrated that the altered protein had abnormal subcellular localization, diminished activity to demethylate the DNA damage-response protein MDC1, and reduced binding to MECP2. We confirmed that JMJD1C protein is widely expressed in brain regions and that its depletion compromises dendritic activity. Conclusions: our findings indicate that mutations in JMJD1C contribute to the development of Rett syndrome and intellectual disability