Epstein-Barr Virus and Human Herpesvirus 6 Detection in a Non-Hodgkin's Diffuse Large B-Cell Lymphoma Cohort by Using RNA Sequencing

Strong, Michael J.; O’Grady, Tina; Lin, Zhen; Xu, Guiyin; Baddoo, Melody; Parsons, Chris; Zhang, Kun; Taylor, Christopher M.; Flemington, Erik K.

doi:10.1128/jvi.02380-13

Cited by 39 publications

(56 citation statements)

References 25 publications

(24 reference statements)

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“…Remarkably, BZLF1 lytic gene expression is suggestive of an abortive lytic cycle linked to tumor progression, as proposed by Strong et al, who recently reported high immediate early lytic expression in DLBCL cases without other lytic genes. 33 A strong correlation among all EBNAs quantified was observed, which is predictable, as in latency III, these EBNA mRNAs are generated by alternative splicing of long primary transcripts initiated either from the tandemly repeated Wp promoter or the upstream Cp promoter. 34 Alternatively, in B cells displaying latency III type infection, LMP1 and LMP2A/B expressions are dependent on expression of EBNA2.…”

Section: Discussionmentioning

confidence: 91%

Epstein-Barr virus-positive diffuse large B-cell lymphoma association is not only restricted to elderly patients

Cohen

Narbaitz²,

Metrebian³

et al. 2014

Int. J. Cancer

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Diffuse large B-cell lymphoma (DLBCL), the most common group of malignant lymphomas, account for 30% of adult nonHodgkin lymphomas. The 2008 World Health Organization (WHO) classification included a new entity, Epstein-Barr virus (EBV)1 DLBCL of the elderly, affecting patients aged 50 years or older. However, some reports of younger EBV1 DLBCL cases, without evidence of underlying immunosuppression, can be found. The role of EBV in tumor microenvironment composition in DLBCL is still not well understood. Our aim was to assess EBV presence and latency pattern as well as tumor T-cell population in an adult DLBCL series of Argentina. The study was conducted on biopsies from 75 DLBCL patients. EBERs expression was performed by in situ hybridization, while EBV gene expression was analyzed using real-time polymerase chain reaction. LMP1, LMP2A, EBNA2, EBNA3A, CD4, CD8 and Foxp3 expression was assessed by immunohistochemistry. Nine percent of cases showed EBV expression, with similar frequency among patients younger than 50 years and 50 years or older (13% and 8%, respectively). T-cell subsets were not altered by EBV presence. Latency type II was the most frequently observed, together with lytic gene expression in EBV1 DLBCL, with 20% of EBERs1 cells. These findings suggest that EBV1 DLBCL in our series was similar to the previously described in Asia and Latin-America, displaying latency II or III expression profile and no agespecific characteristics. Finally, EBV1 DLBCL may be an entity that is not only restricted to patients who are older than 50 years of age, in consequence the age cutoff revision may be a current goal.

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Section: Discussionmentioning

confidence: 91%

Epstein-Barr virus-positive diffuse large B-cell lymphoma association is not only restricted to elderly patients

Cohen

Narbaitz²,

Metrebian³

et al. 2014

Int. J. Cancer

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“…The average number of reads per cell line was approximately 164 million, ranging from 74.6 million to 209.8 million. Based on our previous investigations of microbial agents in biological specimens and cell lines (13)(14)(15), these read depths were deemed to be well above the minimum levels required to detect meaningful pathological infections.…”

Section: Resultsmentioning

confidence: 99%

High-Throughput RNA Sequencing-Based Virome Analysis of 50 Lymphoma Cell Lines from the Cancer Cell Line Encyclopedia Project

Cao¹,

Strong²,

Wang³

et al. 2015

J Virol

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Using high-throughput RNA sequencing data from 50 common lymphoma cell culture models from the Cancer Cell Line Encyclopedia project, we performed an unbiased global interrogation for the presence of a panel of 740 viruses and strains known to infect human and other mammalian cells. This led to the findings of previously identified infections by Epstein-Barr virus (EBV), Kaposi's sarcoma herpesvirus (KSHV), and human T-lymphotropic virus type 1 (HTLV-1). In addition, we also found a previously unreported infection of one cell line (DEL) with a murine leukemia virus. High expression of murine leukemia virus (MuLV) transcripts was observed in DEL cells, and we identified four transcriptionally active integration sites, one being in the TNFRSF6B gene. We also found low levels of MuLV reads in a number of other cell lines and provided evidence suggesting crosscontamination during sequencing. Analysis of HTLV-1 integrations in two cell lines, HuT 102 and MJ, identified 14 and 66 transcriptionally active integration sites with potentially activating integrations in immune regulatory genes, including interleukin-15 (IL-15), IL-6ST, STAT5B, HIVEP1, and IL-9R. Although KSHV and EBV do not typically integrate into the genome, we investigated a previously identified integration of EBV into the BACH2 locus in Raji cells. This analysis identified a BACH2 disruption mechanism involving splice donor sequestration. Through viral gene expression analysis, we detected expression of stable intronic RNAs from the EBV BamHI W repeats that may be part of long transcripts spanning the repeat region. We also observed transcripts at the EBV vIL-10 locus exclusively in the Hodgkin's lymphoma cell line, Hs 611.T, the expression of which were uncoupled from other lytic genes. Assessment of the KSHV viral transcriptome in BCP-1 cells showed expression of the viral immune regulators, K2/vIL-6, K4/vIL-8-like vCCL1, and K5/E2-ubiquitin ligase 1 that was significantly higher than expression of the latency-associated nuclear antigen. Together, this investigation sheds light into the virus composition across these lymphoma model systems and provides insights into common viral mechanistic principles. IMPORTANCEViruses cause cancer in humans. In lymphomas the Epstein-Barr virus (EBV), Kaposi's sarcoma herpesvirus (KSHV) and human T-lymphotropic virus type 1 are major contributors to oncogenesis. We assessed virus-host interactions using a high throughput sequencing method that facilitates the discovery of new virus-host associations and the investigation into how the viruses alter their host environment. We found a previously unknown murine leukemia virus infection in one cell line. We identified cellular genes, including cytokine regulators, that are disrupted by virus integration, and we determined mechanisms through which virus integration causes deregulation of cellular gene expression. Investigation into the KSHV transcriptome in the BCP-1 cell line revealed high-level expression of immune signaling genes. EBV transcriptome analysis sh...

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“…Cancer genome characterization initiatives, including The Cancer Genome Atlas (TCGA), have provided next-generation sequencing (NGS) data on thousands of tumors and various human cancers, and these data are valuable sources for understanding the transcriptional basis of tumorigenesis, as well as variations in tumors (4). Computational techniques have been used to quantify viral presence in many cancers, thereby accelerating the identification of virus-disease associations (5)(6)(7). However, the increasing sensitivity of these techniques may also reveal previously unappreciated viral contaminations that could be erroneously associated with the disease (8)(9)(10).…”

mentioning

confidence: 99%

Possible Human Papillomavirus 38 Contamination of Endometrial Cancer RNA Sequencing Samples in The Cancer Genome Atlas Database

Kazemian

Ren

Lin

et al. 2015

J Virol

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Viruses are causally associated with a number of human malignancies. In this study, we sought to identify new virus-cancer associations by searching RNA sequencing data sets from >2,000 patients, encompassing 21 cancers from The Cancer Genome Atlas (TCGA), for the presence of viral sequences. In agreement with previous studies, we found human papillomavirus 16 (HPV16) and HPV18 in oropharyngeal cancer and hepatitis B and C viruses in liver cancer. Unexpectedly, however, we found HPV38, a cutaneous form of HPV associated with skin cancer, in 32 of 168 samples from endometrial cancer. In 12 of the HPV38-positive (HPV38 ؉ ) samples, we observed at least one paired read that mapped to both human and HPV38 genomes, indicative of viral integration into the host DNA, something not previously demonstrated for HPV38. The expression levels of HPV38 transcripts were relatively low, and all 32 HPV38؉ samples belonged to the same experimental batch of 40 samples, whereas none of the other 128 endometrial carcinoma samples were HPV38 ؉ , raising doubts about the significance of the HPV38 association. Moreover, the HPV38 ؉ samples contained the same 10 novel single nucleotide variations (SNVs), leading us to hypothesize that one patient was infected with this new isolate of HPV38, which was integrated into his/her genome and may have cross-contaminated other TCGA samples within batch 228. Based on our analysis, we propose guidelines to examine the batch effect, virus expression level, and SNVs as part of next-generation sequencing (NGS) data analysis for evaluating the significance of viral/pathogen sequences in clinical samples. IMPORTANCEHigh-throughput RNA sequencing (RNA-Seq), followed by computational analysis, has vastly accelerated the identification of viral and other pathogenic sequences in clinical samples, but cross-contamination during the processing of the samples remain a major problem that can lead to erroneous conclusions. We found HPV38 sequences specifically present in RNA-Seq samples from endometrial cancer patients from TCGA, a virus not previously associated with this type of cancer. However, multiple lines of evidence suggest possible cross-contamination in these samples, which were processed together in the same batch. Despite this potential cross-contamination, our data indicate that we have detected a new isolate of HPV38 that appears to be integrated into the human genome. We also provide general guidelines for computational detection and interpretation of pathogen-disease associations.

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Epstein-Barr Virus and Human Herpesvirus 6 Detection in a Non-Hodgkin's Diffuse Large B-Cell Lymphoma Cohort by Using RNA Sequencing

Cited by 39 publications

References 25 publications

Epstein-Barr virus-positive diffuse large B-cell lymphoma association is not only restricted to elderly patients

Epstein-Barr virus-positive diffuse large B-cell lymphoma association is not only restricted to elderly patients

High-Throughput RNA Sequencing-Based Virome Analysis of 50 Lymphoma Cell Lines from the Cancer Cell Line Encyclopedia Project

Possible Human Papillomavirus 38 Contamination of Endometrial Cancer RNA Sequencing Samples in The Cancer Genome Atlas Database

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