Gaucher Disease: A First Reported Adult Case in Indonesia.

A 44-year-old female presented with a distended abdomen and fatigue. On physical examination, prominent splenomegaly was found. The laboratory investigations revealed pancytopenia and decreased albumin-globulin ratio. The abdominal ultrasonography revealed splenomegaly, cholelithiasis, and cystitis, and the bone survey showed osteopenia. Differential diagnoses included leukemia, multiple myeloma, and myelofibrosis therefore bone marrow puncture was performed. However, histopathologic examination found Gaucher-like cells in the bone marrow aspiration. The finding of CD68 positivity in Gaucher-like cells by using the immunohistochemistry staining supporting Gaucher disease. To confirm the diagnosis, an examination of glucocerebroside substrate from the patient's blood plasma was performed. Glucosylsphingosine, a deacylated form of glucosylceramide, was markedly elevated. Therefore, the diagnosis of Gaucher disease was confirmed. This is the first reported adult Gaucher case diagnosed in Indonesia.

Developing Models to Predict BRAFV600E and RAS Mutational Status in Papillary Thyroid Carcinoma Using Clinicopathological Features and pERK1/2 Immunohistochemistry Expression.

The Cancer Genome Atlas (TCGA) has classified papillary thyroid carcinoma (PTC) into indolent RAS-like and aggressive BRAF-like based on its distinct driver gene mutations. This retrospective study aimed to assess clinicopathology and pERK1/2 expression variations between BRAF-like and RAS-like PTCs and establish predictive models for BRAFV600E and RAS-mutated PTCs. A total of 222 PTCs underwent immunohistochemistry staining to assess pERK1/2 expression and Sanger sequencing to analyze the BRAF and RAS genes. Multivariate logistic regression was employed to develop prediction models. Independent predictors of the BRAFV600E mutation include a nuclear score of 3, the absence of capsules, an aggressive histology subtype, and pERK1/2 levels exceeding 10% (X2 = 0.128, p > 0.05, AUC = 0.734, p < 0.001). The RAS mutation predictive model includes follicular histology subtype and pERK1/2 expression > 10% (X2 = 0.174, p > 0.05, AUC = 0.8, p < 0.001). We propose using the prediction model concurrently with four potential combination group outcomes. PTC cases included in a combination of the low-BRAFV600E-scoring group and high-RAS-scoring group are categorized as RAS-like (adjOR = 4.857, p = 0.01, 95% CI = 1.470-16.049). PTCs included in a combination of the high-BRAFV600E-scoring group and low-RAS-scoring group are categorized as BRAF-like PTCs (adjOR = 3.091, p = 0.001, 95% CI = 1.594-5.995). The different prediction models indicate variations in biological behavior between BRAF-like and RAS-like PTCs.

Profile of BRAFV600E, BRAFK601E, NRAS, HRAS, and KRAS Mutational Status, and Clinicopathological Characteristics of Papillary Thyroid Carcinoma in Indonesian National Referral Hospital.

Introduction: BRAFV600E and RAS mutations are the most common gene mutations in papillary thyroid carcinoma (PTC) that may be correlated with its biological behavior. There are still limited data about BRAFV600E and RAS mutations in Indonesia. This study aims to determine the prevalence of BRAFV600E and RAS mutations, and their association with clinicopathologic characteristics. Methods: Patients who had total thyroidectomy from 2019 to 2021 and those who met our study criteria underwent PCR and DNA sequencing analysis for BRAFV600E, BRAFK601E, exon 2 and 3 of NRAS, HRAS, and KRAS. Analyses were performed to determine the associations of BRAFV600E and RAS mutations with clinicopathologic characteristics. Results: Of 172 PTC patients, BRAFV600E mutation was observed in 37.8% of the patients and RAS mutations were found in 21.5%. One patient harbored BRAFK601E mutation. There was a significant association of BRAFV600E with a high-stage (p = 0.033, OR: 3.279; 95% CI: 1.048-10.259), tall-cell variants (p ≤0.001, OR: 41.143; 95% CI: 11.979-141.308), non-encapsulated (p = 0.001, OR: 4.176; 95% CI: 2.008-8.685), lymphovascular invasion (p = 0.043, OR: 1.912; 95% CI: 1.018-3.592), extrathyroidal extension (p = <0.001, OR: 3.983; 95% CI: 1.970-8.054), and lymph node metastasis (p = 0.009, OR: 2.301; 95% CI: 1.224-4.326). Follicular variant (p = 0.001, OR: 7.011; 95% CI: 2.690-18.268), encapsulated (p = 0.017, OR: 2.433; 95% CI: 1.161-5.100), and absent of extrathyroidal extension (p = 0.033, OR: 2.890; 95% CI: 1.052-7.940) were associated with RAS mutations. Conclusion: A significant association between BRAFV600E mutation and high clinical stage, tall-cell variants, non-encapsulated morphology, lymphovascular invasion, extrathyroidal extension, and lymph node metastasis in PTC was observed. RAS mutations were associated with the follicular variant, encapsulated tumor, and no extrathyroidal extension. HRAS-mutated PTC frequently exhibited tumor multifocality.

Association of Programmed Death-Ligand 1 Expression with Aggressive Histological Types of Thyroid Carcinoma.

Introduction: Immunohistochemical expression of programmed death-ligand 1 (PD-L1) has become a biomarker to predict the usefulness of cancer immunotherapy using PD-1/PD-L1 blockade in a variety of advanced-stage tumours. This emerging biomarker may serve to generate novel therapies for aggressive thyroid carcinoma (TC), which has not shown optimal results with existing treatments. Methods: The present study investigated the relevance of PD-L1 expression in aggressive histological types of TC compared with that found in less aggressive types. Surgically resected specimens were investigated, including 52 cases of TC consisting of 26 cases of aggressive histological types and 26 cases of less aggressive histological types. Immunohistochemical examinations were carried out on paraffin blocks of both groups using a mouse monoclonal primary antibody against PD-L1 (clone 22C3). PD-L1 expression was evaluated by calculating the tumour proportion score (TPS) in both groups. Results: The results revealed a significant difference in the median TPS value of PD-L1 expression between the two groups. The TPS values were found to be higher in the group of aggressive histological types of TC compared with those in the group of less aggressive histological types. A significant difference in TPS value was also found for the extrathyroidal extension variable. Discussion: In conclusion, the present study found a significant association between PD-L1 expression and the aggressive histological type of TC. In addition, a potential association between PD-L1 expression and the presence of extrathyroidal extension of TC was observed. These findings provide novel approaches for immunotherapy as a potential new treatment modality in patients with aggressive histological types of TC.

The Role of Brentuximab Vedotin (BV) as Second-Line Treatment of Refractory Classical Hodgkin Lymphoma (cHL) in the Era of Pandemic COVID-19.

Background: Salvage conventional chemotherapy followed by high-dose therapy (HDT) and autologous stem cell transplantation (ASCT) is the treatment of choice for most patients with refractory classical Hodgkin lymphoma (cHL). In the era of pandemic COVID-19, there are obstacles to administering salvage chemotherapy followed by HDT and ASCT due to side effects and toxicities which make the patient more susceptible to COVID-19 infection. The toxicities and side effects of BV are different from salvage chemotherapy, but it has clear efficacy as monotherapy. Case Presentation: A 46-year-old female with a history of cHL nodular sclerosis subtype was presented with right cervical lymph node enlargement, after 3 cycles of first-line chemotherapy ABVD (adriamycin, bleomycin, vinblastine, and dacarbazine) 3 months ago. She was afraid to undergo second-line chemotherapy in the era of pandemic COVID-19 because of the side effects and toxicities; therefore, she was given 8 cycles of BV as monotherapy. The response of the treatment was complete remission. Conclusion: In this particular case of patient, BV as monotherapy can be an option during the pandemic COVID-19 for refractory cHL ineligible for second-line chemotherapy followed by HDT and ASCT.

MUM1 Expression versus Hans Algorithm to Predict Prognosis in Indonesian Diffuse Large B-Cell Lymphoma Patients Receiving R-CHOP.

Background: Treatment response in diffuse large B-cell lymphoma (DLBCL) is heterogenous. The Hans algorithm (using 30% cut-offs for CD10, BCL6, and MUM1 protein expression) has been the most favored method to categorize DLBCL into germinal center B-cell (GCB) and non-GCB subtypes in order to predict prognosis. However, the algorithm's ability to prognosticate is not always consistent. Methods: This retrospective cohort study was conducted on DLBCL patients receiving R-CHOP therapy at Dr. Cipto Mangunkusumo Hospital, Jakarta from 2014 to 2017. We aimed to compare the prognostic value of Hans algorithm as well as the protein levels of CD10, BCL6, MUM1, and Ki67 at different cut-offs. Ninety-two patients were classified based on Hans algorithm and various proteins at different cut-off values were analyzed with regard to event-free survival at 24 months using survival analysis. The cut-off values were then compared using receiver operating characteristic curves. Results: A significant survival difference was observed with MUM1 expression cut-off of 50% or more (log rank p = 0.035). CD10, BCL6, Ki67, and Hans algorithm showed AUCs below or near 0.5 (0.405, 0.436, 0.498, and 0.413, respectively), whereas MUM1 showed an AUC of 0.835, in predicting events within 24 months. MUM-1 cut-off of 70.5% yielded an optimal trade-off for sensitivity and specificity. Conclusion: MUM1 expression of 50% or more can help predict prognosis in DLBCL patients receiving R-CHOP therapy and can be considered as for use as a single marker to predict prognosis.

Clinicopathological Profile of Thyroid Carcinoma in Young Patients: An Indonesian Single-Center Study.

INTRODUCTION: Thyroid cancer is the third most common cancer that occurs in children and adolescents. Papillary thyroid carcinoma (PTC) is the most common type of thyroid malignancy. Although the mortality rate of thyroid malignancy in children is usually low, the disease recurrence is higher in children with more severe clinical presentation than in adults. This study aimed to determine the demographic and clinicopathological characteristics and outcome of pediatric and adolescent patients with thyroid malignancy in Indonesia. METHODS: The retrospective study included all patients diagnosed with thyroid carcinoma aged <20 years, from January 1, 2015, to December 31, 2019. Twenty-nine subjects fulfilled the inclusion and exclusion criteria. We retrieved baseline characteristics, pathology features, TSH and fT4 status, radioactive iodine therapy data, and patients' outcomes. Then, data were analyzed using the chi-square or Fisher's exact method. RESULTS: We identified 29 eligible subjects, including 3 boys and 26 girls. The most common type of thyroid carcinoma was PTC (96.5%), and follicular type (31%) was the predominant variant of PTC. Lymph node involvement occurred in 24% of patients, while distant metastasis occurred in 17.2% of patients with PTC. Twenty-four (82.7%) patients had stage 1 disease. Disease recurrence was recorded in 31% of patients during the study period with a median follow-up time of 24 months. CONCLUSION: PTC is the most frequent type of thyroid carcinoma among children and adolescents. This malignancy has a low mortality rate, but the recurrence rate remains high among younger patients than adults even during a short-term follow-up analysis. Distant metastasis and lymph node involvement are commonly found in this age group.

Expression of programmed cell death ligand-1 protein in germinal center B-cell-like and non-germinal center B-cell-like subtypes of diffuse large B-cell lymphoma.

Diffuse large B-cell lymphoma (DLBCL) is the most common type of non-Hodgkin lymphoma worldwide. However, there is still limited information on the expression of programmed cell death ligand-1 (PD-L1), a type 1 transmembrane protein of immunoglobulin B7/CD28 in the DLBCL subtypes. The present study aimed to identify the expression of PD-L1 in germinal center B-cell-like (GCB) subtype and non-germinal center B-cell-like (non-GCB) subtype of DLBCL. A total of 40 patient samples (formalin-fixed paraffin-embedded tissues) consisting of 20 cases of GCB subtype and 20 cases of non-GCB subtype of DLBCL were examined. The PD-L1 protein expressions were evaluated by using immunohistochemical staining in the tumor cells. The results showed a statistically significant difference (P=0.003) between the expression of PD-L1 in the GCB subtype and the non-GCB subtype of DLBCL. PD-L1 expression in the tumor cells were observed in 13 cases (65%) of non-GCB subtype and in three cases (15%) of the GCB subtype of DLBCL. In conclusion, it was found that the expression of PD-L1 protein in the tumor cells of the non-GCB subtype of DLBCL was significantly higher as compared with the tumor cells of the GCB subtype of DLBCL.

Less than subtotal parathyroidectomy in multiple endocrine neoplasia type 1: A case report and review of the literature.

INTRODUCTION: Multiple endocrine neoplasia type 1 (MEN1) is a rare syndrome with common presenting signs and symptoms secondary to primary hyperparathyroidism (PHPT), which is managed surgically. Traditionally, either subtotal parathyroidectomy (SPTX) or total parathyroidectomy with autologous transplantation (TPTX) is the approach of choice. However, recent studies showed comparable persistence and recurrence rate in a subset of MEN1 patients (two or more concordant preoperative imaging results) who underwent less than subtotal parathyroidectomy (LSPTX). PRESENTATION OF CASE: We report a case of patient with PHPT and delayed diagnosis of MEN1, who underwent LSPTX without intraoperative parathyroid hormone (IOPTH) measurement. The approach was chosen based on the preoperative imaging studies. Unfortunately, the PHPT persisted and the patient was reoperated. To further elucidate the issue, a systematic search of the literature was conducted on Cochrane library, PubMed, and Scopus; articles relevant to the case were reviewed. Results are conflicting results with most of the studies showed LSPTX is inferior compared to the other two approaches. DISCUSSION: Therefore, given the current body of evidence, we consider that subtotal or total parathyroidectomy is still the preferred surgical approach for the treatment of PHPT in MEN1. CONCLUSION: Further studies are still needed to see whether LSPTX is comparable to SPTX or TPTX in regards to persistent and recurrent hyperparathyroidism if the conditions are met.

Natural killer cell neoplasm: biology and pathology.

Natural killer (NK) cell neoplasm is a heterogeneous disease group. In the latest World Health Organization (WHO) classification of tumours of hematopoietic and lymphoid tissues (2008), disease entities considered as NK-cell derivation include NK-lymphoblastic leukemia/lymphoma, chronic lymphoproliferative disorders of NK cells, aggressive NK-cell leukemia, and extranodal NK-cell lymphoma, nasal-type. Despite recent advances in NK-cell research, which have expanded our understanding of the biology of NK-cell neoplasm, it cannot yet be sharply delineated from myeloid neoplasms and T-cell neoplasms even in some "well-known" entity, such as extranodal NK/T-cell lymphoma. This review describes current knowledge of the biology of NK cells and pathology of NK neoplasms as classified in the 2008 WHO classification of tumours of hematopoietic and lymphoid tissues.

Malignant lymphomas in Waldeyer's ring in Asian countries: Association with histologic types and Epstein-Barr virus.

The present study examines clinicopathologic findings and their association with the Epstein-Barr virus (EBV) in Waldeyer's ring lymphomas (WRLs) from Indonesia (91 cases), P.R. China (31 cases), Korea (101 cases) and Japan (61 cases). Waldeyer's ring (WR) was categorized into upper and lower parts comprising the pharyngeal and tubal tonsils (upper WR) or palatine and lingual tonsils (lower WR), respectively. Diffuse large B-cell lymphoma (DLBCL) pre-dominated in the lower WR in all countries at a frequency of 78.9-100%. Natural killer/T-cell lymphoma (NKTCL) was predominant in the upper WR in China, Korea and Japan at a frequency of 50-62.5%, while in Indonesia it occcurred at a frequency of less than 10%. On the whole, patients with NKTCL were significantly younger (median 43 years) than those with DLBCL (57 years). Patients with DLBCL in the lower WR were significantly younger in Indonesia (median 50 years) than in China (63 years) or Japan (69 years). The percentage of EBV-positive cases was much higher in NKTCL (78.6-100%) than in DLBCL (2.2-6%). This study evaluates the differences between East and Southeast Asian countries in terms of histologic type and age distribution in WRLs categorized by the location of the lesions in WR.

Condensin mutations and abnormal chromosomal structures in pyothorax-associated lymphoma.

Transfer of genetic information during mitosis is accurately conducted by proper condensation and segregation of chromosomes, for which condensins play a central role. Both condensin I and II have common structural maintenance of chromosomes subunits, named hCAP-C and hCAP-E. Pyothorax-associated lymphoma (PAL) is a non-Hodgkin's lymphoma developing in the pleural cavity of patients with long-standing pyothorax. Mutations of hCAP-C and hCAP-E were investigated in 24 leukemia-lymphoma cell lines including eight PAL cell lines, and their influences in chromosome morphology were evaluated. Heterozygous point mutations within hCAP-C were found in two PAL cell lines and corresponding tumor samples (OPL-3 and OPL-7). Deletion of exon 24 within hCAP-E and a point mutation at the donor splice site of intron 24 were detected in OPL-5 and original tumor samples. OPL-5 showed an extensive reduction in expression of not only hCAP-E but also hCAP-C proteins. OPL-5 occasionally showed the chromosome bridge in anaphase and telophase, indicating that segregation is not accurate. OPL-7 showed reduced hCAP-C protein expression, abnormality in chromosome length and width, and abnormal aggregates of hCAP-C protein. These findings indicated that condensin gene alteration might play a role in genome instability, which accelerates the accumulation of other gene alterations in PAL.

Telomere dysfunction and inactivation of the p16(INK4a)/Rb pathway in pyothorax-associated lymphoma.

Previous studies have indicated that genome instability is involved in the lymphomagenesis of pyothorax-associated lymphoma (PAL), which develops in patients with a long-standing history of pyothorax. One of the well-known causes of genome instability is telomere dysfunction. In the present study, the condition of telomeres was analyzed in the cell lines and clinical samples from PAL. Telomere length (TL) in PAL cell lines was extremely short (<4.5 kbp). TL in tumor samples was broad in range, and shorter than that in the peripheral blood leukocytes from the matched patients. Three of five PAL cell lines showed frequent loss of telomere signals (telomere erosion); however, telomerase activity in PAL cell lines was similar to that in Burkitt lymphoma cell lines. Rb expression was detected in three PAL cell lines and four of 15 clinical samples, respectively. Rb protein expressed in three PAL cell lines was heavily phosphorylated, indicating that function of Rb protein was suppressed. p16(INK4a) expression was not detected in either cell lines or clinical samples. The promoter region in p16(INK4a) was heavily methylated in all cell lines as well as the clinical samples. Inactivation of the p16(INK4a)/Rb pathway may allow continuous cell division and critical telomere shortening, which induce genome instability, finally leading to malignant transformation. Taken together, telomere dysfunction and inactivation of the p16(INK4a)/Rb pathway might play a role for PAL development.

Gene mutation analysis of sinonasal lymphomas in Indonesia.

Sinonasal lymphomas comprise NK/T-cell (NKTCL) type and B-cell type with unique geographical development. In this study, mutations of p53, K-ras, c-kit, beta-catenin, and bak gene were analyzed using polymerase chain reaction (PCR)-single strand conformation polymorphism (SSCP) followed by direct sequencing in 41 sinonasal lymphomas (27 NKTCL and 14 B-cell type) from Indonesia. In situ hybridization study with EBER-1 probe revealed that 85% of NKTCL cases were EBV positive, but none of B-cell type was EBV positive. Frequency of mutations in p53, K-ras, c-kit, beta-catenin, and bak gene was 62.9%, 0%, 11.1%, 18.5%, and 25.9%, respectively, in NKTCL, and 71.4%, 0%, 23.1%, 21.4%, and 57.1%, respectively, in B-cell cases, showing that mutation frequency in all genes was higher in B-cell than in NKTCL cases. These findings suggest that gene mutations might be the driving-force for B-cell lymphoma, whereas combined EBV infection and gene mutations contribute to NKTCL development in Indonesia.

Impairment of double-strand breaks repair and aberrant splicing of ATM and MRE11 in leukemia-lymphoma cell lines with microsatellite instability.

Mutations of DNA double-strand breaks (DSB) repair genes, ATM, MRE11, RAD50, NBS1 and ATR, are postulated to play a role in the development of gastrointestinal malignancies with an impaired mismatch repair (MMR) function. In the present study, mutations of these genes together with the presence of microsatellite instability (MSI) were examined in 50 leukemia-lymphoma cell lines. MSI was detected in 13 (26%) lines. Mutations of intronic mononucleotide repeats in ATM and MRE11 were found in nine and six lines, respectively, whereas mutations of mononucleotide repeats of RAD50 were found in only one line, and none were found in either NBS1 or ATR. Frequencies of ATM and MRE11 mutations were significantly higher in MSI-positive than MSI-negative lines. These mutations generated aberrant splicing in both genes. The intensity of the aberrant transcript of ATM (497del22) was stronger in five lines harboring mononucleotide mutations of 2 bp or more than in the lines without or with a 1-bp mutation. The intensity of the aberrant transcript of MRE11 (315del88) was stronger in four lines with mononucleotide mutations than in lines without. The expression levels of ATM and MRE11 transcripts in MSI-positive lines were significantly higher than those in MSI-negative lines. MSI-positive cell lines showed delay or abrogation of DSB repair. The present study suggests that impairment of the MMR system causes aberrant transcripts in the DSB repair genes ATM and MRE11. This might result in inactivation of the DSB repair system, thus inducing an acceleration of genome instability and accumulation of genetic damage.

Loss of expression of Epstein-Barr virus nuclear antigen-2 correlates with a poor prognosis in cases of pyothorax-associated lymphoma.

Pyothorax-associated lymphoma (PAL) is a non-Hodgkin's lymphoma, which develops in the pleural cavity of patients who have had pyothorax for over 20 years, and is strongly associated with Epstein-Barr virus (EBV) infection. The expression of latent genes, especially EBV nuclear antigen-2 (EBNA-2), influences the growth characteristics and malignant features of EBV-infected cells. Here, the effect of EBNA-2 expression on clinical features was examined in 13 cases of PAL. The EBNA-2 transcript was detected in 8 cases but was absent in 5. There was a significant difference in survival between patients with the transcripts and those without: the 1-year survival rate was 87.5 and 0%, respectively (p < 0.01). There was a discrepancy between EBNA-2 expression and EBNA promoter usage in 6 cases. The Cp/Wp promoter was used in 3 EBNA-2-negative cases, whereas the Qp promoter or multiple promoters were used in 3 EBNA-2-positive cases. Analysis of PAL cell lines provided a clue as to the mechanism underlying the discrepancy between EBNA-2 expression and EBNA promoter usage. Loss of EBNA-2 expression, irrespective of the latency pattern, is correlated with a poor prognosis, suggesting that down-regulation of the EBNA-2 expression could be selection pressure for the progression of PAL.

Alterations of DNA damage-response genes ATM and ATR in pyothorax-associated lymphoma.

Pyothorax-associated lymphoma (PAL) is non-Hodgkin's lymphoma that develops from chronic inflammation. Free radicals and oxidative stress generated in the inflammatory lesions could cause DNA damage and thus provide a basis for lymphomagenesis. Ataxia-telangiectasia mutated (ATM) and ATM and Rad3-related (ATR) genes are responsive genes for DNA damage, therefore potential involvement of these genes in PAL lymphomagenesis was examined in eight PAL cell lines and clinical samples from five cases. ATM mutations were detected in five of eight PAL lines. All but one of these mutations affected the phosphatidylinositol 3-kinase domain, indicating the loss-of-function mutation of ATM gene. Heterozygous mutations of ATR were found in two of eight lines; one a missense and the other a truncation mutation. ATR mutations were also detected in two of five cases in clinical samples from PAL. PAL cells with ATR mutation showed a delay or abrogation in repair for ionizing radiation (IR)-induced DNA double-strand breaks (DSBs) or ultraviolet (UV)-induced DNA single-strand breaks (SSBs), and exhibited a defect in p53 accumulation and failure in cell cycle checkpoint at G1-S phase. These findings showed that mutations of ATR gene result in failure for DNA DSB and SSB repair, suggesting the role of ATM and ATR gene mutations in PAL lymphomagenesis.

Identification of Epstein-Barr virus integrated sites in lymphoblastoid cell line (IB4).

IB4 is a lymphoblastoid cell line frequently used for the functional analysis of the latent genes of EBV. Previous study indicated that EBV whole genome is integrated tandemly as the linear viral genome into host genome of IB4, although sites of integration have not been determined. Through cloning of the junctional regions between EBV and host genomes, one of the integration sites was identified on the BamHI-C fragments around oriP sequences and another on the EcoRI-I fragment. Both of the integration sites were located on the clone RP11-119H12 of chromosome 4q25 and separated approximately 6.5 kbp from each other. The integration sites identified were apart from the genes of the host genome, indicating that both host gene and EBV latent genes are not altered by the integration event.

Epstein-Barr virus is integrated between REL and BCL-11A in American Burkitt lymphoma cell line (NAB-2).

Epstein-Barr virus (EBV) initially isolated from the cultured Burkitt lymphoma (BL) cells, is one of the well-known oncogenic virus. The NAB-2 line, which was established from a North American Burkitt's tumor, was indicated to contain one copy of EBV DNA as the integrated form into chromosome 2p13 of the host genome. To demonstrate the integration site of EBV directly, and to clarify the relation between the integration sites and the oncogenes, fragments containing the nucleotide sequence of NAB-2 integration sites were cloned. EBV was integrated via the terminal repeats (TR), and integration sites located in the clone RP11-440P5 on chromosome 2, between two oncogenes, REL and BCL11A, which is apart from approximately 350 kbp from each other. Expression level of REL in NAB-2 was increased. The flanking region of chromosome 2 at the bilateral junction sites showed no homology to the junction sites of EBV. The integration site 2p13 overlaps with common fragile site, FRA2E. NAB-2 cells expressed almost all latent genes but LMP-2A that flanks the TR, indicating the type III of latent infection of EBV. Integration event in NAB-2 might alter the regulation of the oncogenes and provide advantage for continuous cell proliferation.