Although clonal selection by genetic driver aberrations in cancer is well documented, the ability of epigenetic alterations to promote tumor evolution is undefined. We used 450k arrays and ...next-generation sequencing to evaluate intratumor heterogeneity and evolution of DNA methylation and genetic aberrations in chronic lymphocytic leukemia (CLL). CLL cases exhibit vast interpatient differences in intratumor methylation heterogeneity, with genetically clonal cases maintaining low methylation heterogeneity and up to 10% of total CpGs in a monoallelically methylated state. Increasing methylation heterogeneity correlates with advanced genetic subclonal complexity. Selection of novel DNA methylation patterns is observed only in cases that undergo genetic evolution, and independent genetic evolution is uncommon and is restricted to low-risk alterations. These results reveal that although evolution of DNA methylation occurs in high-risk, clinically progressive cases, positive selection of novel methylation patterns entails coevolution of genetic alteration(s) in CLL.
Understanding the molecular and phenotypic heterogeneity of cancer is a prerequisite for effective treatment. For chronic lymphocytic leukemia (CLL), recurrent genetic driver events have been ...extensively cataloged, but this does not suffice to explain the disease's diverse course. Here, we performed RNA-sequencing on 184 CLL patient samples. Unsupervised analysis revealed two major, orthogonal axes of gene expression variation: the first one represented the mutational status of the immunoglobulin heavy variable (IGHV) genes, and concomitantly, the three-group stratification of CLL by global DNA methylation. The second axis aligned with trisomy 12 status and affected chemokine, MAPK and mTOR signaling. We discovered nonadditive effects (epistasis) of IGHV mutation status and trisomy 12 on multiple phenotypes, including the expression of 893 genes. Multiple types of epistasis were observed, including synergy, buffering, suppression and inversion, suggesting that molecular understanding of disease heterogeneity requires studying such genetic events not only individually but in combination. We detected strong differentially expressed gene signatures associated with major gene mutations and copy-number aberrations including SF3B1, BRAF and TP53, as well as del(17)(p13), del(13)(q14) and del(11)(q22.3) beyond dosage effect. Our study reveals previously underappreciated gene expression signatures for the major molecular subtypes in CLL and the presence of epistasis between them.
Novel agents such as the Bcl-2 inhibitor venetoclax (ABT-199) are changing treatment paradigms for chronic lymphocytic leukemia (CLL) but important problems remain. Although some patients exhibit ...deep and durable responses to venetoclax as a single agent, other patients harbor subpopulations of resistant leukemia cells that mediate disease recurrence. One hypothesis for the origin of resistance to venetoclax is by kinase-mediated survival signals encountered in proliferation centers that may be unique for individual patients. An in vitro microenvironment model was developed with primary CLL cells that could be incorporated into an automated high-content microscopy-based screen of kinase inhibitors (KIs) to identify agents that may improve venetoclax therapy in a personalized manner. Marked interpatient variability was noted for which KIs were effective; nevertheless, sunitinib was identified as the most common clinically available KI effective in overcoming venetoclax resistance. Examination of the underlying mechanisms indicated that venetoclax resistance may be induced by microenvironmental signals that upregulate antiapoptotic Bcl-xl, Mcl-1, and A1, which can be counteracted more efficiently by sunitinib than by ibrutinib or idelalisib. Although patient-specific drug responses are common, for many patients, combination therapy with sunitinib may significantly improve the therapeutic efficacy of venetoclax.
•Patient-specific pathways of resistance to venetoclax can be identified by high-content screening of clinical samples with a KI library.•Sunitinib may overcome resistance to venetoclax for many patients by downregulating the expression of Bcl-xl, Mcl-1, and A1 in CLL cells.
Abstract
Background
The 2 cofactors in the etiology of Burkitt lymphoma (BL) are Epstein-Barr virus (EBV) and repeated Plasmodium falciparum malaria infections. This study evaluated EBV loads in ...mucosal and systemic compartments of children with malaria and controls. Age was analyzed as a covariate because immunity to malaria in endemic regions is age dependent.
Methods
Children (2–10 years) with clinical malaria from Western Kenya and community controls without malaria were enrolled. Saliva and blood samples were collected, EBV viral load was assessed by quantitative polymerase chain reaction, and EpiTYPER MassARRAY was used to assess methylation of 3 different EBV genes.
Results
Regardless of the compartment, we detected EBV more frequently in malaria cases compared to controls, although the difference was not significant. When EBV was detected, there were no differences in viral load between cases and controls. However, EBV methylation was significantly lower in the malaria group compared to controls in both plasma and saliva (P < .05), indicating increased EBV lytic replication. In younger children before development of immunity to malaria, there was a significant effect of malaria on EBV load in peripheral blood mononuclear cells (P = .04).
Conclusions
These data suggest that malaria can directly modulate EBV persistence in children, increasing their risk for BL.
In this study, we determined that concurrent malaria infection results in higher lytic EBV DNA in saliva and plasma of children, indicative of malaria-induced viral reactivation and higher frequency of EBV-infected cells in PBMCS that is age dependent.
Acute myeloid leukemia (AML) is a molecularly complex disease characterized by heterogeneous tumor genetic profiles and involving numerous pathogenic mechanisms and pathways. Integration of molecular ...data types across multiple patient cohorts may advance current genetic approaches for improved subclassification and understanding of the biology of the disease. Here, we analyzed genome-wide DNA methylation in 649 AML patients using Illumina arrays and identified a configuration of 13 subtypes (termed "epitypes") using unbiased clustering. Integration of genetic data revealed that most epitypes were associated with a certain recurrent mutation (or combination) in a majority of patients, yet other epitypes were largely independent. Epitypes showed developmental blockage at discrete stages of myeloid differentiation, revealing epitypes that retain arrested hematopoietic stem-cell-like phenotypes. Detailed analyses of DNA methylation patterns identified unique patterns of aberrant hyper- and hypomethylation among epitypes, with variable involvement of transcription factors influencing promoter, enhancer, and repressed regions. Patients in epitypes with stem-cell-like methylation features showed inferior overall survival along with up-regulated stem cell gene expression signatures. We further identified a DNA methylation signature involving STAT motifs associated with
-ITD mutations. Finally, DNA methylation signatures were stable at relapse for the large majority of patients, and rare epitype switching accompanied loss of the dominant epitype mutations and reversion to stem-cell-like methylation patterns. These results show that DNA methylation-based classification integrates important molecular features of AML to reveal the diverse pathogenic and biological aspects of the disease.
Prostate cancer is driven by a combination of genetic and/or epigenetic alterations. Epigenetic alterations are frequently observed in all human cancers, yet how aberrant epigenetic signatures are ...established is poorly understood. Here we show that the gene encoding BAZ2A (TIP5), a factor previously implicated in epigenetic rRNA gene silencing, is overexpressed in prostate cancer and is paradoxically involved in maintaining prostate cancer cell growth, a feature specific to cancer cells. BAZ2A regulates numerous protein-coding genes and directly interacts with EZH2 to maintain epigenetic silencing at genes repressed in metastasis. BAZ2A overexpression is tightly associated with a molecular subtype displaying a CpG island methylator phenotype (CIMP). Finally, high BAZ2A levels serve as an independent predictor of biochemical recurrence in a cohort of 7,682 individuals with prostate cancer. This work identifies a new aberrant role for the epigenetic regulator BAZ2A, which can also serve as a useful marker for metastatic potential in prostate cancer.
CD21low age-associated or atypical memory B cells are autoantibody enriched and poised for plasma cell differentiation. These cells overaccumulate in chronic infections, autoimmune disease, and ...immunodeficiency, posing the question of what checkpoints normally oppose their accumulation. Here, we reveal a critical role for paralogous calcium-NFAT-regulated transcription factors EGR2 and EGR3 that are induced in self-reactive B cells. CD21low and B1 B cells lacking EGR2 and EGR3 accumulate and circulate in young mice in numbers 10- to 20-fold greater than normal and overexpress a large set of EGR2 ChIP-seq target genes, including known drivers of plasma cell differentiation. Most follicular B cells constitutively express Egr2 proportionally to surface IgM downregulation by self-antigens, and EGR2/3 deficiency abolishes this cardinal feature of B cell anergy. These results explain the cardinal features of B cell anergy, define a key transcriptional checkpoint repressing CD21low B cell formation, and inform how NFATC1 or EGR2 mutations promote B1 cell-derived chronic lymphocytic leukemias.
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•Follicular B cells express Egr2 proportionally to IgM cell-surface downregulation•Egr2/3 deficiency decreases Zfp318 and increases surface IgM expression•EGR2 and EGR3 are transcriptional regulators of atypical CD21low B cells•EGR2 and EGR3 prevent excessive accumulation of CD21low B cells and B1 cells
Chronic B cell receptor signaling induces the transcription factors EGR2/3. EGR2 mutations recur in chronic lymphocytic leukemia. Masle-Farquhar et al. show that EGR2/3 regulate the cardinal features of B cell anergy and prevent the overaccumulation of mouse B1 lymphocytes and atypical CD21low B lymphocytes resembling those that accumulate with age, chronic infections, immunodeficiency, and autoimmunity
Despite much evidence on epigenetic abnormalities in cancer, it is currently unclear to what extent epigenetic alterations can be associated with tumors’ clonal genetic origins. Here, we show that ...the prostate intratumor heterogeneity in DNA methylation and copy-number patterns can be explained by a unified evolutionary process. By assaying multiple topographically distinct tumor sites, premalignant lesions, and lymph node metastases within five cases of prostate cancer, we demonstrate that both DNA methylation and copy-number heterogeneity consistently reflect the life history of the tumors. Furthermore, we show cases of genetic or epigenetic convergent evolution and highlight the diversity in the evolutionary origins and aberration spectrum between tumor and metastatic subclones. Importantly, DNA methylation can complement genetic data by serving as a proxy for activity at regulatory domains, as we show through identification of high epigenetic heterogeneity at androgen-receptor-bound enhancers. Epigenome variation thereby expands on the current genome-centric view on tumor heterogeneity.
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•Epigenetic intratumor heterogeneity reflects prostate tumors’ clonal genetic origins•Tumor and metastatic subclones exhibit diverse evolutionary origins•Distinct tumor lineages show convergent evolution of methylation patterns•Degree of methylation variability at a locus depends on genetic and epigenetic context
Several recent studies have examined the clonal origins of tumor populations on the basis of genetic variability. Brocks et al. now show that subclonal lineages also differ in their DNA methylation patterns and that methylation heterogeneity is consistent with the tumors’ genetic history. Furthermore, high methylation variability at enhancers bound by androgen receptor, which are known to play a role in prostate tumorigenesis, suggests that DNA methylation data can serve as a proxy for the functional state of tumor subclones.
Radiotherapy is a fundamental part of cancer treatment but its use is limited by the onset of late adverse effects in the normal tissue, especially radiation-induced fibrosis. Since the molecular ...causes for fibrosis are largely unknown, we analyse if epigenetic regulation might explain inter-individual differences in fibrosis risk. DNA methylation profiling of dermal fibroblasts obtained from breast cancer patients prior to irradiation identifies differences associated with fibrosis. One region is characterized as a differentially methylated enhancer of diacylglycerol kinase alpha (DGKA). Decreased DNA methylation at this enhancer enables recruitment of the profibrotic transcription factor early growth response 1 (EGR1) and facilitates radiation-induced DGKA transcription in cells from patients later developing fibrosis. Conversely, inhibition of DGKA has pronounced effects on diacylglycerol-mediated lipid homeostasis and reduces profibrotic fibroblast activation. Collectively, DGKA is an epigenetically deregulated kinase involved in radiation response and may serve as a marker and therapeutic target for personalized radiotherapy.
Recently, the European LeukemiaNet (ELN) revised its genetic-risk classification of acute myeloid leukemia (AML). We categorized 1637 adults with AML treated with cytarabine/anthracycline regimens ...according to the 2022 and 2017 ELN classifications. Compared with the 2017 ELN classification, 2022 favorable group decreased from 40% to 35% and adverse group increased from 37% to 41% of patients. The 2022 genetic-risk groups seemed to accurately reflect treatment outcomes in all patients and patients aged <60 years, but in patients aged ≥60 years, relapse rates, disease-free (DFS) and overall (OS) survival were not significantly different between intermediate and adverse groups. In younger African-American patients, DFS and OS did not differ between intermediate-risk and adverse-risk patients nor did DFS between favorable and intermediate groups. In Hispanic patients, DFS and OS did not differ between favorable and intermediate groups. Outcome prediction abilities of 2022 and 2017 ELN classifications were similar. Among favorable-risk patients, myelodysplasia-related mutations did not affect patients with CEBPA
mutations or core-binding factor AML, but changed risk assignment of NPM1-mutated/FLT3-ITD-negative patients to intermediate. NPM1-mutated patients with adverse-risk cytogenetic abnormalities were closer prognostically to the intermediate than adverse group. Our analyses both confirm and challenge prognostic significance of some of the newly added markers.
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