Signaling pathways of EBV-induced oncogenesis.

Epstein-Barr virus (EBV) is closely associated with multiple human cancers. EBV-associated cancers are mainly lymphomas derived from B cells and T cells (Hodgkin lymphoma, Burkitt lymphoma, NK/T-cell lymphoma, and posttransplant lymphoproliferative disorder (PTLD)) and carcinomas derived from epithelial cells (nasopharyngeal carcinoma and gastric carcinoma). EBV can induce oncogenesis in its host cell by activating various signaling pathways, such as nuclear factor-κB (NF-κB), phosphoinositide-3-kinase/protein kinase B (PI3K/AKT), Janus kinase/signal transducer and transcription activator (JAK/STAT), mitogen-activated protein kinase (MAPK), transforming growth factor-ß (TGF-ß), and Wnt/ß-catenin, which are regulated by EBV-encoded proteins and noncoding RNA. In this review, we focus on the oncogenic roles of EBV that are mediated through the aforementioned signaling pathways.

Molecular mechanisms of EBV-driven cell cycle progression and oncogenesis.

The early stage of oncogenesis is linked to the disorder of the cell cycle. Abnormal gene expression often leads to cell cycle disorders, resulting in malignant transformation of human cells. Epstein-Barr virus (EBV) is associated with a diverse range of human neoplasms, such as malignant lymphoma, nasopharyngeal carcinoma and gastric cancer. EBV mainly infects human lymphocytes and oropharyngeal epithelial cells. EBV is latent in lymphocytes for a long period of time, is detached from the cytoplasm by circular DNA, and can integrate into the chromosome of cells. EBV expresses a variety of latent genes during latent infection. The interaction between EBV latent genes and oncogenes leads to host cell cycle disturbances, including the promotion of G1/S phase transition and inhibition of cell apoptosis, thereby promoting the development of EBV-associated neoplasms. Molecular mechanisms of EBV-driven cell cycle progression and oncogenesis involve diverse genes and signal pathways. Here, we review the molecular mechanisms of EBV-driven cell cycle progression and promoting oncogenesis.

[MiR-503 sensitizes human hepatocellular carcinoma cells to cisplatin by targeting bcl-2].

OBJECTIVE: To investigate effects of miR-503 on cisplatin sensitivity in BEL-7402 cells by targeting of bcl-2.
 Methods: MiR-503 and bcl-2 mRNA expression levels in hepatocellular carcinoma cells were measured by real-time quantitative (qRT)-PCR; Bcl-protein level was detected by Western blot; miR-503 mimics were transiently transfected to the BEL-7402 cells by liposome transfection; potential target genes of miR-503 were predicted by Bioinformatics software; miR-503 potential targets were validated by dual luciferase activity; and the cell viability was measured by MTT assay. 
 Results: MiR-503 level was down-regulated and Bcl-2 protein expression level was up-regulated in BEL-7402 cells compared with HL-7702 cells. MiR-503 could interact with bcl-2 and inhibit its expression. Cell vitality with miR-503 transfection was significantly reduced compared to that in the negative control.
 Conclusion: MiR-503 may enhance the sensitivity of BEL-7402 cells to cisplatin and inhibit the cell proliferation by targeting bcl-2.

Identification of IgH gene rearrangement and immunophenotype in an animal model of Epstein-Barr virus-associated lymphomas.

Epstein-Barr virus (EBV) is a human oncogenic herpesvirus associated with lymphoma and nasopharyngeal carcinoma. Because the susceptible hosts of EB virus are limited to human and cotton-top tamarins (Saguinus oedipus), there have been no appropriate animal models until the lymphoma model induced by EBV in human peripheral blood lymphocyte (hu-PBL)/SCID chimeric mice was reported. However, it is still controversial whether the EBV-associated lymphoma induced in hu-PBL/SCID mice is a monoclonal tumor. In this study, we transplanted normal human peripheral blood lymphocytes (hu-PBL) from six donors infected with EBV into SCID mice to construct hu-PBL/SCID chimeric mice. The induced tumors were found in the mediastinum or abdominal cavity of SCID mice. Microscopic observation exhibited tumor cells that were large and had a plasmablastic, centroblastic or immunoblastic-like appearance. Immunophenotyping assays showed the induced tumors were LCA-positive, CD20/CD79a-positive (markers of B cells), and CD3/CD45RO-negative (markers of T cells). A human-specific Alu sequence could be amplified by Alu-PCR. This confirmed that induced tumors were B-cell lymphomas originating from the transplanted human lymphocytes rather than mouse cells. EBER in situ hybridization detected positive signals in the nuclei of the tumor cells. Expression of EBV-encoded LMP1, EBNA-1, and EBNA-2 in the tumors was significantly positive. PCR-based capillary electrophoresis analysis of IgH gene rearrangement revealed a monoclonal peak and single amplification product in all six cases of induced tumors. This indicated that EBV can induce monoclonal proliferation of human B lymphocytes and promotes the development of lymphoma. J. Med. Virol. 88:1804-1813, 2016. © 2016 Wiley Periodicals, Inc.

Wnt signaling in gastric cancer: current progress and future prospects.

Levels of the Wnt pathway components are abnormally altered in gastric cancer cells, leading to malignant cell proliferation, invasion and metastasis, poor prognosis and chemoresistance. Therefore, it is important to understand the mechanism of Wnt signaling pathway in gastric cancer. We systematically reviewed the molecular mechanisms of the Wnt pathway in gastric cancer development; and summarize the progression and the challenges of research on molecular agents of the Wnt pathway.

Screening and functional analysis of differentially expressed genes in EBV-transformed lymphoblasts.

BACKGROUND: Epstain-Barr virus (EBV) can transform human B lymphocytes making them immortalized and inducing tumorigenic ability in vitro, but the molecular mechanisms remain unclear. The aim of the present study is to detect and analyze differentially expressed genes in two types of host cells, normal human lymphocytes and coupled EBV-transformed lymphoblasts in vitro using gene chips, and to screen the key regulatory genes of lymphocyte transformation induced by EB virus. METHODS: Fresh peripheral blood samples from seven healthy donors were collected. EBV was used to transform lymphocytes in vitro. Total RNA was extracted from 7 cases of the normal lymphocytes and transformed lymphoblasts respectively, marked with dihydroxyfluorane after reverse transcription, then hybridized with 4 × 44 K Agilent human whole genome microarray. LIMMA, String, Cytoscape and other softwares were used to screen and analyze differentially expressed genes. Real-time PCR was applied to verify the result of gene expression microarrays. RESULTS: There were 1745 differentially expressed genes that had been screened, including 917 up-regulated genes and 828 down-regulated genes. According to the results of Generank, String and Cytoscape analyses, 38 genes may be key controlled genes related to EBV-transformed lymphocytes, including 22 up-regulated genes(PLK1, E2F1, AURKB, CDK2, PLCG2, CD80, PIK3R3, CDC20, CDC6, AURKA, CENPA, BUB1B, NUP37, MAD2L1, BIRC5, CDC25A, CCNB1, RPA3, HJURP, KIF2C, CDK1, CDCA8) and 16 down-regulated genes(FYN, CD3D, CD4, CD3G, ZAP70, FOS, HCK, CD247, PRKCQ, ITK, LCP2, CXCL1, CD8A, ITGB5, VAV3, CXCR4), which primarily control biological processes such as cell cycle, mitosis, cytokine-cytokine pathway, immunity response and so on. CONCLUSIONS: Human lymphocyte transformation induced by EB virus is a complicated process, involving multiple-genes and -pathways in virus-host interactions. Global gene expression profile analysis showed that EBV may transform human B lymphocytes by promoting cell cycle and mitosis, inhibiting cell apoptosis, hindering host immune function and secretion of cytokines.

The Wnt/ß-catenin signalling pathway in Haematological Neoplasms.

Leukaemia and lymphoma are common malignancies. The Wnt pathway is a complex network of proteins regulating cell proliferation and differentiation, as well as cancer development, and is divided into the Wnt/ß-catenin signalling pathway (the canonical Wnt signalling pathway) and the noncanonical Wnt signalling pathway. The Wnt/ß-catenin signalling pathway is highly conserved evolutionarily, and activation or inhibition of either of the pathways may lead to cancer development and progression. The aim of this review is to analyse the mechanisms of action of related molecules in the Wnt/ß-catenin pathway in haematologic malignancies and their feasibility as therapeutic targets.

miR-4295 Promotes the Malignant Progression of Gastric Cancer via Targeting PTEN.

BACKGROUND: Gastric cancer (GC), one of the common clinical malignant tumors of the digestive system, is the fourth most commonly diagnosed cancer and the second lethal cancer worldwide and has the characteristics of high metastasis, fatality, and recurrence rate. This research was conducted to investigate the role and mechanism of miR-4295 in gastric cancer. METHODS: The expression capacity of miR-4295 was determined in gastric cancer tissues and its normal tissues by qRT-PCR. PTEN expression level was detected by western blot. SGC-7901 and MGC-803 cell lines were cultured and transfected with miR-4295 or its inhibitor. The effects of miR-4295 on cell proliferation, colony formation, migration, and invasion in vitro were investigated. The mutual effect between miR-4295 and PTEN in 293T cells was explored by luciferase reporter gene assays. RESULTS: The results showed that miR-4295 expression was higher in gastric cancer tissues and cell lines, and the miR-4295 level was significantly negatively associated with the tumor size and distal metastasis of gastric cancer. Notably, up-regulated miR-4295 promoted cell proliferation, migration and invasion in vitro, whereas it led to contrary effects while down-regulating miR-4295 expression. Further mechanism studies displayed that miR-4295 could directly fasten the PTEN 3'UTR and dramatically decrease the level of PTEN in vitro. CONCLUSION: The findings revealed that miR-4295 could promote gastric cancer cell proliferation, migration and invasion, which might be attributed to targeting PTEN. Our study suggested that miR-4295 might be a potential therapeutic target for gastric cancer.

Human-derived IgG level as an indicator for EBV-associated lymphoma model in Hu-PBL/SCID chimeras.

BACKGROUND: Epstein-Barr virus (EBV) has a close association with various types of human lymphomas. Animal models are essential to elucidate the pathogenesis of human EBV-associated lymphomas. The aim of the present study is to evaluate the association between human IgG concentration and EBV-associated lymphoma development in huPBL/SCID mice. METHODS: Human peripheral blood lymphocytes (hu-PBL) from EBV-seropositive donors were inoculated intraperitoneally into SCID mouse. Immunohistochemical staining was used to examine differentiated antigens of tumor cells. EBV infection of the induced tumors was detected by in situ hybridization. IgG concentrations in the serums of 12 SCID mice were measured by unidirectional immunodiffusion assay. RESULTS: 21 out of 29 mice developed tumors in their body. Immunohistochemical staining showed that all induced tumors were LCA (leukocyte common antigen) positive, B-cell markers (CD20, CD79a) positive, and T-cell markers (both CD3 and CD45RO) negative. The tumors can be diagnosed as human B-cell lymphomas by these morphological and immunohistochemical features. In situ hybridization exhibited resultant tumor cells had EBV encoded small RNA-1 (EBER-1). Human-derived IgG could be found in the serum from SCID mice on the 15th day following hu-PBL transplantation, and IgG levels increased with the tumor development in 6 hu-PBL/SCID chimeras. CONCLUSIONS: Intraperitoneal transfer of hu-PBLs from EBV+ donors to SCID mice leads to high human IgG levels in mouse serum and B cell lymphomas. Our findings suggest that increasing levels of human-derived IgG in peripheral blood from hu-PBL/SCID mice could be used to monitor EBV-related human B-cell lymphoma development in experimental animals.

Clinical crosstalk between microRNAs and gastric cancer (Review).

Globally, there were over 1 million new gastric cancer (GC) patients in 2018 and GC has become the sixth most common cancer worldwide. GC caused 783,000 deaths worldwide in 2018, making it the third most deadly cancer type. miRNAs are short (~22 nucleotides in length) non­coding RNA molecules, which can regulate gene expression passively at a post­transcriptional level. There are more and more in­depth studies on miRNAs. There are numerous conclusive evidences that there is an inseparable link between miRNAs and GC. miRNAs can affect the entire process of GC, including the oncogenesis, development, diagnosis, treatment and prognosis of GC. Although many miRNAs have been linked to GC, few can be applied to clinical practice. This review takes the clinical changes of GC as a clue and summarizes the miRNAs related to GC that have confirmed the mechanism of action in the past three years. Through in­depth study and understanding of the mechanism of those miRNAs, we predict their possible clinical uses, and suggest some new insights to overcome GC.

Gene analysis of Epstein-Barr virus-associated lymphomas in Hu-Pbl/SCID chimeras.

AIMS AND BACKGROUND: The mechanisms of Epstein-Barr virus (EBV)-associated tumor development are incompletely understood. The aim of this study was to investigate the gene expression of EBV-associated lymphomas in hu-PBL/SCID mice. METHODS: Human peripheral blood lymphocytes (hu-PBL) from EBV-seropositive donors were transplanted into severe combined immunodeficiency (SCID) mice. In situ hybridization was used to detect EBV-encoded small RNA- 1 (EBER1) in tumor tissues. Mutation of TP53 exons 5-8 in EBV-induced lymphomas was analyzed by PCR-SSCP. Immunohistochemical staining was used to examine EBV gene products and cellular oncoproteins. RESULTS: Twenty-one of 29 mice developed tumors. EBER1 was positive in the nuclei of almost all tumor cells. Immunohistochemistry showed positive staining of LMP1, EBNA2 and ZEBRA in a small number of tumor cells. Immunohistochemically detectable p53 protein expression was common (85.7%), but TP53 gene mutations were identified in only four cases (19.1%) of EBV-associated lymphomas. Positivity rates of C-myc, Bcl-2 and Bax expression were 100%, 95.2%, and 90.5%, respectively, in the 21 cases of EBV-associated lymphomas. CONCLUSIONS: Our preliminary findings suggest that EBV-associated lymphomas in hu-PBL/SCID chimeras show EBV infection, expression of oncogenic viral genes, and overexpression of cellular oncogenes. TP53 gene mutations are rare but p53 protein is commonly expressed in EBV-associated lymphomas.

Long non-coding RNA GAS5 in human cancer.

Long non-coding RNAs (lncRNAs) constitute a group of >200-nucleotide ncRNA molecules. lncRNAs regulate several cell functions, such as proliferation, apoptosis, invasion and metastasis. Meanwhile, lncRNAs are abnormally expressed in human malignancies, where they suppress or promote tumor growth. The present study focused on growth arrest-specific transcript 5 (GAS5), a well-known lncRNA that acts as a tumor suppressor but is suppressed in multiple types of cancer, including mammary carcinoma, prostate cancer, colorectal cancer, gastric cancer, melanoma, esophageal squamous cell carcinoma, lung cancer, ovarian cancer, cervical cancer, gliomas, osteosarcoma, pancreatic cancer, bladder cancer, kidney cancer, papillary thyroid carcinoma, neuroblastoma, endometrial cancer and liver cancer. Notably, GAS5 is overexpressed in liver cancer, potentially functioning as an oncogene. In the present study, the diagnostic and therapeutic roles of GAS5 in different tumors were reviewed, with a summary of the potential clinical application of the lncRNA, which may help identify novel study directions for GAS5.

Epstein-Barr virus-associated gastric cancer: A distinct subtype.

Epstein-Barr virus (EBV)-associated gastric cancer (EBVaGC) is a common malignant tumor associated with EBV infection. The molecular classification of gastric carcinoma indicates that EBVaGC is a distinct subtype in terms of oncogenesis and molecular features. Viral proteins, Bam-HI-A rightward transcripts (BART) miRNAs, and Bam-HI A rightward frame 1 (BARF1) promote oncogenesis after EBV infection via the induction of methylation, regulation of host gene expression, and malignant transformation. Together with abnormal mutations and amplification of the host genome as driving factors, interactions between the EBV genome and host genome accelerate carcinogenesis. The molecular profile of EBVaGC is that of EBV driving DNA hypermethylation, frequent phosphatidylinositol-4,5-bisphosphate 3-kinase, catalytic subunit alpha (PIK3CA) mutations, and the overexpression of Janus kinase 2 (JAK2), programmed death ligand-1 (PD-L1), and PD-L2. Clinically, the frequency of lymph node metastasis is lower, and the prognosis is better for EBVaGC than EBV-negative gastric cancer (EBVnGC). Pathologically, EBVaGC is a gastric adenocarcinoma with lymphoid stroma. This review interprets how the EBV genome is involved in the oncogenesis of gastric cancer and describes the molecular and clinicopathological features of EBVaGC.

Identification of prognostic aging-related genes associated with immunosuppression and inflammation in head and neck squamous cell carcinoma.

Aging is regarded as a dominant risk factor for cancer. Additionally, inflammation and asthenic immune surveillance with aging may facilitate tumor formation and development. However, few studies have comprehensively analyzed the relationship between aging-related genes (AGs) and the prognosis, inflammation and tumor immunity of head and neck squamous cell carcinoma (HNSCC). Here, we initially screened 41 differentially expressed AGs from The Cancer Genome Atlas (TCGA) database. In the training set, a prognosis risk model with seven AGs (APP, CDKN2A, EGFR, HSPD1, IL2RG, PLAU and VEGFA) was constructed and validated in the TCGA test set and the GEO set (P < 0.05). Using univariate and multivariate Cox regression analyses, we confirmed that risk score was an independent prognostic factor of HNSCC patients. In addition, a high risk score was significantly correlated with immunosuppression, and high expression of PLAU, APP and EGFR was the main factor. Furthermore, we confirmed that a high risk score was significantly associated with levels of proinflammatory factors (IL-1α, IL-1ß, IL-6 and IL-8) in HNSCC samples. Thus, this risk model may serve as a prognostic signature and provide clues for individualized immunotherapy for HNSCC patients.

Bioinformatic analysis of differentially expressed genes and identification of key genes in EBV-transformed lymphoblasts.

Although the Epstein-Barr virus (EBV) is a well-known human oncogenic virus, its molecular mechanisms involved in the transformation of healthy human cells remain poorly understood. In this study, human lymphocytes were isolated from the peripheral blood of healthy adults, and lymphocytes were transformed in vitro by EBV. Agilent human whole genome microarrays were used to detect the differential gene expression profiles of EBV-transformed lymphoblasts and healthy peripheral blood lymphocytes (PBLs). By constructing the gene functional network of EBV-induced lymphocyte transformation, we screened out candidate key genes in this process and verified their expression levels by real-time quantitative polymerase chain reaction (RT-qPCR) and Western blot. In the EBV-transformed lymphoblasts, 2335 differentially expressed genes, including 1328 up-regulated and 1007 down-regulated, were screened out. Five candidate key genes, namely, PLK1, E2F1, PTPN11, BIRC5 and FYN were mainly screened out according to the results of LIMMA, String, Cytoscape software analysis. RT-qPCR and Western blot showed that PLK1, E2F1, PTPN11, BIRC5 genes had increased expression levels, and FYN gene was down-regulated in EBV-transformed lymphoblasts. Silencing of PLK1 gene in Raji cells could inhibit cell proliferation and invasion, and induce cell cycle arrest and apoptosis. In conclusion, PLK1, E2F1, PTPN11, BIRC5 and FYN are the candidate key molecules of EBV-transformed lymphocytes.

MiR-33b-5p sensitizes gastric cancer cells to chemotherapy drugs via inhibiting HMGA2 expression.

MicroRNAs (miRNAs) are internal, non-coding, and ∼22 nt small RNAs that display cell- and tissue-specific expression. They play important regulatory roles in cell proliferation and chemo-sensitivity. This study focused on tumor-suppressive miR-33b-5p expression as well as its role in gastric cancer. MiR-33b-5p was found low expression in gastric cancer cell lines. Functionally, western blots and the luciferase reporter assay were used to confirm that HMGA2 was the potential target of miR-33b-5p. Next, we used CCK-8 kits to analyze the effect of miR-33b-5p combined chemotherapy drugs on cell inhibition rate, and flow cytometry to analyze cells apoptosis. Colony formation ability was determined by plating at 500 cells per well into six-well plates and culturing for 15 d. The results showed that upregulation of miR-33b-5p decreased expression of HMGA2 and inhibited gastric cancer cell growth as well as sensitized gastric cancer cells to chemotherapy drugs. MiR-33b-5p overexpression hindered luciferase activity of HMGA2,3'-untranslated region-based reporter construct in 293 T cells. These data demonstrate that miR-33b-5p may be a potential therapeutic target for gastric cancer and function as tumor-suppressive miRNA through targeting HMGA2 in gastric cancer.

miR-503 inhibits proliferation making human hepatocellular carcinoma cells susceptible to 5­fluorouracil by targeting EIF4E.

Hepatocellular carcinoma (HCC), a disease that is a major health care issue across the globe, includes the deviant expression of miRNAs in its development, progression, and resistance to treatment. We focused our study on miR­503 expression and its role in HCC. miR­503 was found in HCC tissues and cell lines using quantitative real-time PCR (RT­qPCR). Western blot analyses and the luciferase reporter assay were used to determine the miR­503 potential target in the HCC cells. We used MTT to analyze cell proliferation activity and noted that there was a considerable decrease of miR­503 in HCC tissues and cell lines when measured against the controls. miR­503 upregulation decreased expression of eukaryotic translation initiation factor 4E (EIF4E), and reduced HCC cell proliferation and sensitized HCC cells to anticancer drugs. miR­503 overexpression hindered luciferase activity of EIF4E 3' untranslated region-based reporter construct among HepG2, BEL-7402, and SMMC-7721 cells, revealing that miR­503 may increase sensitivity to therapies at least partially through targeting EIF4E suppression of HCC proliferation.

Detection and location of Helicobacter pylori in human gastric carcinomas.

AIM: To define the infection status of Helicobacter pylori in 109 patients with gastric cancers and H pylori localization in gastric carcinoma tissues in South China. METHODS: The incidence of H pylori infection in gastric carcinomas was estimated by polymerase chain reaction (PCR), simultaneously; both morphological features and the localization of H pylori in gastric carcinomas were demonstrated by Warthin-Starry (WS) staining. The relationships between H pylori infection and the clinical-pathologic factors of gastric carcinomas were analyzed by software SPSS10.0. RESULTS: H pylori was found in 42 (39.03%) and 58 (53.21%) cases of 109 patients with gastric carcinomas by PCR and WS, respectively. H pylori infection rate detected in gastric carcinomas by WS was higher than that by PCR (chi2 = 9.735, P < 0.005 < 0.01). WS stain showed that H pylori existed in the gastric antrum mucus, mucosal gland of normal tissues adjacent to gastric carcinomas and the gland, mucus pool of cancer tissues. The positive rate of H pylori in normal tissues adjacent to carcinomas was higher than that in cancer tissues (chi2 = 15.750, P < 0.005 < 0.01). No significant differences in age, sex, site, histological types and lymph node metastasis were found between H pylori-positive gastric carcinomas and H pylori-negative cases by both methods, but there were statistically significant differences of H pylori positive rate between early and advanced stage of gastric carcinomas (chi2 = 4.548 or 5.922, P = 0.033 or 0.015 < 0.05). CONCLUSION: These results suggested that H pylori infection might play a certain role in the early stage of carcinogenesis of human gastric mucosa epithelia.

Construction of hu-PBL/SCID chimeras and development of EBV-related lymphomas.

OBJECTIVE: To construct hu-PBL/SCID chimeras and to investigate the development of lymphoma and oncogenicity of the Epstein-Barr virus (EBV). METHODS: Human peripheral blood lymphocytes (PBLs) were isolated from healthy adult donors and transplanted intraperitoneally into severe combined immunodeficient (SCID) mice. Mice with hu-PBL engraftment from healthy EBV seronegative donors were injected intraperitoneally with EBV-containing supernatant from suspension culture of B95-8 cell line (active infection), whereas mice receiving lymphocytes from healthy EBV seropositive donors were not re-infected with B95-8 derived EBV (latent infection). Pathological examination and molecular analysis were performed on experimental animals and induced neoplasms. RESULTS: In the early stage of this experiment, 12 mice died of acute graft-versus-host disease, mortality was 34.3% (12/35 mice) with an average life span of 17.5 days. In 19 survival hu-PBL/SCID chimeric recipients from 12 healthy donors, tumor incidence was 84.2% (16/19 mice). The average survival time of tumor-bearing mice was 65.5 days. EBV-related neoplasms in SCID mice were nodular tumors with aggressive and fatal features. Histological morphology of tumors exhibited diffuse large cell lymphomas. Immunohistochemistry revealed that LCA (CD45) and L26 (CD20) were positive, but both PS1 (CD3) and UCHL-1 (CD45RO) were negative, and EBV products ZEBRA, LMP1, and EBNA2 were expressed in a small number of tumor cells. EB virus particles were seen in the nuclei of some tumor cells by electron microscopy, and EBV DNA could be amplified in the tumor tissues by PCR. In situ hybridization indicated that the nuclei of tumor cells contained human-specific Alu sequence. CONCLUSIONS: EBV-induced tumors were human B-cell malignant lymphomas. We obtained direct causative evidence dealing with EBV-associated tumor deriving from normal human cells.