Glioblastoma with novel EGFR mutations (T790M and exon 20 insertion) yet unresponsive to osimertinib: A case report.

Glioblastoma (GBM) is a high-grade adult-type IDH-wildtype diffuse glioma, commonly harboring epidermal growth factor receptor (EGFR) amplification. Here, we describe a case of a 49-year-old man with a GBM harboring a TERT promoter mutation. Despite surgical and chemoradiation therapy, the tumor recurred. At that time, comprehensive genomic profiling by next-generation sequencing identified two rare mutations in EGFR: T790M and an exon 20 insertion. Based on these findings, the patient elected to undergo off-label therapy with osimertinib, a third-generation EGFR tyrosine kinase inhibitor that has shown promising results in non-small cell lung carcinoma, including metastatic to brain, with exactly the same EGFR mutations. Moreover, the drug has excellent central nervous system penetration. Even so, no clinical response was observed, and the patient succumbed to the disease. The lack of response may be related to the specific nature of the EGFR mutations, and/or other unfavorable tumor biology overriding any benefit from osimertinib.

Placenta-Derived Extracellular Vesicles in Pregnancy Complications and Prospects on a Liquid Biopsy for Hemoglobin Bart's Disease.

Extracellular vesicles (EVs) are nano-scaled vesicles released from all cell types into extracellular fluids and specifically contain signature molecules of the original cells and tissues, including the placenta. Placenta-derived EVs can be detected in maternal circulation at as early as six weeks of gestation, and their release can be triggered by the oxygen level and glucose concentration. Placental-associated complications such as preeclampsia, fetal growth restriction, and gestational diabetes have alterations in placenta-derived EVs in maternal plasma, and this can be used as a liquid biopsy for the diagnosis, prediction, and monitoring of such pregnancy complications. Alpha-thalassemia major ("homozygous alpha-thalassemia-1") or hemoglobin Bart's disease is the most severe form of thalassemia disease, and this condition is lethal for the fetus. Women with Bart's hydrops fetalis demonstrate signs of placental hypoxia and placentomegaly, thereby placenta-derived EVs provide an opportunity for a non-invasive liquid biopsy of this lethal condition. In this article, we introduced clinical features and current diagnostic markers of Bart's hydrops fetalis, extensively summarize the characteristics and biology of placenta-derived EVs, and discuss the challenges and opportunities of placenta-derived EVs as part of diagnostic tests for placental complications focusing on Bart's hydrop fetalis.

Excessive Subchorionic Fibrinoid Deposition as a Component of Massive Perivillous Fibrin Deposition: A Case With Maternal Immune Thrombocytopenia.

Maternal floor infarction (MFI) and massive perivillous fibrin deposition (MPFD) are overlapping placental disorders of unknown etiology, associated with adverse obstetric outcome, and a significant risk of recurrence. We describe a 31-year-old mother with asymptomatic thrombocytopenia throughout pregnancy and a positive lupus anticoagulant. She delivered a normal female neonate at term, whose weight was small for gestational age, with a placenta weighing less than the 10th percentile. Placental examination showed MPFD together with excessive subchorionic fibrinoid deposition. The placenta showed diffuse C4d deposition and an immune-mediated reaction was postulated for the pathogenesis of the placental changes. We suggest that excessive subchorionic fibrinoid deposition may be part of the morphologic spectrum of MFI/MPFD.

Pediatric fibromyxoid tumor with PLAG1 fusion: An emerging entity with a novel intracranial location.

Translocations involving PLAG1 occur in several tumors, most commonly pleomorphic adenoma and lipoblastoma. Recently, a distinctive soft tissue tumor with a PLAG1 fusion has been reported in the pediatric age group. These are low grade tumors with a fibroblastic or mixed fibroblastic and myxoid morphology but no other lines of differentiation. They are typically immunopositive for desmin and CD34. The partner genes for these tumors have included YWHAZ, EEF1A1, ZFHX4l, CHCHD7, and PCMTD1. We report another case of this fibromyxoid tumor with a PLAG1 fusion, this time with COL3A1 as the partner gene. The fusion placed expression of a full-length PLAG1 protein under the control of the constitutively active COL3A1 promoter. Overexpression of PLAG1 was confirmed by diffusely positive immunostaining for PLAG1. The most novel aspect of this tumor is the intracranial location. Opinion has been divided over whether these tumors are a specific entity, or related to lipoblastoma, since that tumor also typically occurs in soft tissue in the pediatric age group and shows many of the same gene fusions. However, lipoblastoma has never been reported in an intracranial location and, thus, our case provides compelling evidence that this fibromyxoid tumor is indeed a distinct entity.

Classification of stillbirth by the International Classification of Diseases for Perinatal Mortality using a sequential approach: A 20-year retrospective study from Thailand.

AIM: The International Classification of Diseases for Perinatal Mortality (ICD-PM) is a system for recording causes of perinatal death. In this system, placental pathology is considered a "maternal condition" and this category does not cover the spectrum of placental pathology that can impact on perinatal death. The aim of the study was to apply a wider spectrum of placental pathology as a separate parameter for classifying death in the ICD-PM. METHODS: All autopsy reports at a single institution over a 20-year period (2001-2020) were reviewed. Causes of stillbirth were analyzed in a sequential manner: step 1, clinical history and laboratory results; step 2, placenta; and step 3, autopsy; and classified at each step according to the ICD-PM. RESULTS: The review identified 330 cases, including 126 antepartum and 204 intrapartum deaths. Step 1 identified a cause in 176 (86%) intrapartum deaths and 64 (51%) antepartum deaths. The addition of placental pathology (step 2) changed the cause of death in 12% of cases, with causes now identified in 190 (93%) intrapartum and 89 (71%) antepartum deaths. Adding step 3 did not identify any additional causes of death. CONCLUSION: The accuracy of the ICD-PM classification is dependent on the data available. Placental pathology made a significant difference in assigning causes of death in our series, stressing the importance of placental examination. Determination of the cause of death based on clinical history and laboratory data alone may be inaccurate, and less useful for comparative studies and planning prenatal care.

Pediatric fibromyxoid soft tissue tumor with PLAG1 fusion: A novel entity?

The classification of undifferentiated soft tissue tumors continues to evolve with the expanded application of molecular analysis in clinical practice. We report three cases of a unique soft tissue tumor in young children (5 months to 2 years old) displaying a purely fibromyxoid histology, with positive staining for desmin and CD34. In two cases, RNA sequencing detected a YWHAZ-PLAG1 gene fusion, while in the third case, a previously unreported EEF1A1-PLAG1 fusion was identified. PLAG1 fusions have been reported in several pathologic entities including pleomorphic adenoma, myoepithelial tumors of skin and soft tissue, and lipoblastoma, the latter occurring preferentially in young children. In these tumors, expression of a full length PLAG1 protein comes under the control of the constitutively active promoter of the partner gene in the fusion, and the current cases conform to that model. Overexpression of PLAG1 was confirmed by diffusely positive immunostaining for PLAG1 in all three cases. Our findings raise the possibility of a novel fibromyxoid neoplasm in childhood associated with these rare PLAG1 fusion variants. The only other report of a PLAG1-YWHAZ fusion occurred in a pediatric tumor diagnosed as a "fibroblastic lipoblastoma." This finding raises the possibility of a relationship with our three cases, even though our cases lacked any fat component. Further studies with regard to a shared pathogenesis are required.

Placental Findings Contributing to Perinatal Death: A 15-Year Retrospective Review from a Teaching Hospital in Thailand.

Introduction: The placenta is infrequently examined in developing countries. This study examined the role of placental pathology in perinatal deaths at Chulalongkorn University Hospital, Bangkok. Methods: Included were singleton intrauterine deaths after gestational week 20 and live-born infants up to 1 week old, over a 15-year period. Placental lesions were classified as: inflammatory-immune, maternal stromal-vascular, fetal stromal-vascular, umbilical cord complications and other. Results: 208 such cases had the placenta available. A placental cause of death was found in 96 (46%), non-placental causes in 28% and the cause of death was unknown in 26%. Of those 96 placentas, 44% were categorized as inflammatory-immune, 30% maternal stromal-vascular, 13% fetal stromal-vascular, 7% umbilical cord complications and 6% other. Conclusions: Placental causes of death were less common than in many Western studies, but inflammatory-immune processes more common. These differences may relate to how cases were accrued, and/or local socioeconomic factors, and warrant further study.

Combined Placental Maternal Floor Infarction and Cytomegalovirus Placentitis: A Case Report.

BackgroundMaternal floor infarction (MFI) and massive perivillous fibrin deposition (MPFD) are uncommon, related placental conditions secondary to trophoblastic cell damage. The etiology is unknown but MPFD/MFI is associated with adverse obstetric outcome and a significant risk of recurrence. Case report: We report a case of MPFD/MFI associated with cytomegalovirus (CMV) placentitis. A 27-year-old mother delivered a stillborn male fetus with a postmortem diagnosis of congenital CMV. The placenta showed a lymphohistiocytic villitis with isolated CMV inclusions, in combination with MFI. The villitis had features intermediate between CMV placentitis and villitis of unknown etiology (VUE). Conclusion: VUE is considered to be a maternal anti-fetal immune reaction resembling allograft rejection. We postulate that the viral infection in our case may have triggered this immune response, given that CMV antigens are known to cross react with some human antigens, in particular HLA. The subsequent trophoblastic cell damage could then lead to MFI/MFPD.

Molecular characterization of DICER1-mutated pituitary blastoma.

Pituitary blastoma (PitB) has recently been identified as a rare and potentially lethal pediatric intracranial tumor. All cases that have been studied molecularly possess at least one DICER1 pathogenic variant. Here, we characterized nine pituitary samples, including three fresh frozen PitBs, three normal fetal pituitary glands and three normal postnatal pituitary glands using small-RNA-Seq, RNA-Seq, methylation profiling, whole genome sequencing and Nanostring® miRNA analyses; an extended series of 21 pituitary samples was used for validation purposes. These analyses demonstrated that DICER1 RNase IIIb hotspot mutations in PitBs induced improper processing of miRNA precursors, resulting in aberrant 5p-derived miRNA products and a skewed distribution of miRNAs favoring mature 3p over 5p miRNAs. This led to dysregulation of hundreds of 5p and 3p miRNAs and concomitant dysregulation of numerous mRNA targets. Gene expression analysis revealed PRAME as the most significantly upregulated gene (500-fold increase). PRAME is a member of the Retinoic Acid Receptor (RAR) signaling pathway and in PitBs, the RAR, WNT and NOTCH pathways are dysregulated. Cancer Hallmarks analysis showed that PI3K pathway is activated in the tumors. Whole genome sequencing demonstrated a quiet genome with very few somatic alterations. The comparison of methylation profiles to publicly available data from ~ 3000 other central nervous system tumors revealed that PitBs have a distinct methylation profile compared to all other tumors, including pituitary adenomas. In conclusion, this comprehensive characterization of DICER1-related PitB revealed key molecular underpinnings of PitB and identified pathways that could potentially be exploited in the treatment of this tumor.

Massive Perivillous Fibrin Deposition Associated With Placental Syphilis: A Case Report.

Massive perivillous fibrin deposition (MPFD) and the related entity of maternal floor infarction (MFI) are uncommon placental disorders of unknown etiology, associated with adverse obstetric outcome and a significant risk of recurrence. We describe a 19-year-old mother with untreated syphilis who delivered a male neonate with low birth weight, skin desquamation, and pneumonia. Placenta examination showed the expected changes for syphilis but unexpectedly, also showed MPFD. To our knowledge, this is the first report of MPFD associated with placental syphilis, thus expanding the list of etiologies that may be related to MPFD/MFI. It is postulated that the syphilis infection in our case led to a hypercoaguable state, eventually resulting in MPFD. In the right clinical setting, syphilis might be considered in the differential diagnosis when MPFD/MFI is observed on placental examination. The recurrence risk of MFPD/MFI associated with infections is believed to be lower than idiopathic cases and, by extrapolation, this lower risk should apply to syphilis as well.

Adult-onset neuroblastoma: Report of seven cases with molecular genetic characterization.

Whereas neuroblastoma is the most common extracranial solid tumor of childhood, less than 5% of cases occur in adults. Pediatric neuroblastoma shows marked heterogeneity of histology and molecular biology. Information about this tumor in adults is limited, especially regarding molecular biology. We report a series of nine neuroblastoma cases diagnosed in adulthood (18 to 40 years old) with molecular biologic characterization in seven. All tumors were Schwannian stroma-poor, and mostly poorly differentiated. Tumors expressed neural markers including PHOX2B, NB84, synaptophysin, chromogranin, CD56, neuron-specific enolase, and PGP9.5. Two out of six cases expressed ALK and one had the F1174 L mutation reported in childhood neuroblastoma. Fluorescent in situ hybridization (FISH) revealed MYCN amplification in 2/7 cases, chromosome 1p deletion in 1/5 cases and 17q gain in 4/4 cases. One in five cases showed loss of ATRX expression by immunohistochemistry and alternate lengthening of telomeres by FISH. Zero out of five cases showed rearrangement of the TERT gene by FISH, but one case showed high level amplification. In conclusion, the morphology and immunophenotype of adult-onset neuroblastoma are similar to pediatric cases although less differentiated than some childhood tumors. Similarly, molecular genetic alterations in adult-onset neuroblastoma are not unique to this age group. However, 80% of cases tested showed genetic changes that would promote maintenance of telomeres, which is a molecular marker of high risk cases. This may help explain the poor response in adults to pediatric treatment protocols. Additional studies to characterize the biology of this tumor in the adult age group will facilitate the design of more personalized therapeutic approaches.

Prevalence and Clinical Correlations of Somatostatin Receptor-2 (SSTR2) Expression in Neuroblastoma.

Alternative radiolabeled, targeted agents are being investigated for children with relapsed neuroblastoma (NB) who do not respond to I-metaiodobenzylguanidine (MIBG) therapy. (DOTA-Tyr)-octreotate targets somatostatin receptors (SSTRs), particularly SSTR2, which are expressed on NB cells. We investigated SSTR2 expression in NB tumors (36 high-risk [HR]; 33 non-HR patients) and correlated SSTR2 levels with clinical features, norepinephrine transporter (NET) expression, and MIBG avidity. SSTR2 and NET immunohistochemistry scores (0 to 3) were calculated on biopsies using digital image analysis based on staining intensity and distribution. Clinical data were correlated with SSTR2 expression. Median SSTR2 score for 69 patients was 1.31 (0.26 to 2.55). Non-HR NB was associated with a higher SSTR2 score (P=0.032). The SSTR2 expression did not correlate with age, International Neuroblastoma Staging System (INSS) stage, MYCN amplification and histology. Higher SSTR2 scores were observed in MIBG-avid versus MIBG-nonavid NB. SSTR2 score was not significantly associated with NET score (r=-0.062, P=0.62). Twenty-six patients who relapsed or progressed had a median SSTR2 score of 1.33 (0.26 to 2.55). Patients with NB including relapsed or progressive disease showed SSTR2 expression at diagnosis, suggesting they could be candidates for radiolabeled-DOTA-conjugated peptide imaging or therapy.

Electron Microscopy Can Still Have a Role in the Diagnosis of Selected Inborn Errors of Metabolism.

Many anatomic pathology laboratories no longer have electron microscopy facilities. A retrospective review of autopsies was performed to identify cases of inborn errors of metabolism (IEM) and determine the contribution of electron microscopy in making the diagnosis in those cases. Over a period of 17 years, there were 900 perinatal and pediatric autopsies. There were 7 cases (1%) of IEM, including 4 cases of Pompe disease, 1 case of I-cell disease, 1 case of bile acid synthesis defect, and 1 case of mitochondrial disease (Leigh syndrome). Electron microscopy was important in the diagnosis of I-cell disease and Pompe disease in our series. This technique enabled a prenatal diagnosis to be made from a chorionic villus biopsy in 2 cases with a positive family history. In less developed countries where upfront genetic testing may be too expensive and may need international referral, electron microscopy can still be useful for diagnosis of IEM, providing an affordable alternative with a more rapid turnaround time compared to gene mutation analysis or enzyme assay. Results can be used both for patient management and as a screen for which cases might benefit from genetic testing.

Sarcoma Subgrouping by Detection of Fusion Transcripts Using NanoString nCounter Technology.

BACKGROUND: NanoString technology is an innovative barcode-based system that requires less tissue than traditional techniques and can test for multiple fusion transcripts in a single reaction. The objective of this study was to determine the utility of NanoString technology in the detection of sarcoma-specific fusion transcripts in pediatric sarcomas. DESIGN: Probe pairs for the most common pediatric sarcoma fusion transcripts were designed for the assay. The NanoString assay was used to test 22 specific fusion transcripts in 45 sarcoma samples that had exhibited one of these fusion genes previously by reverse transcription polymerase chain reaction (RT-PCR). A mixture of frozen (n = 18), formalin-fixed, paraffin-embedded (FFPE) tissue (n = 23), and rapid extract template (n = 4) were used for testing. RESULTS: Each of the 22 transcripts tested was detected in at least one of the 45 tumor samples. The results of the NanoString assay were 100% concordant with the previous RT-PCR results for the tumor samples, and the technique was successful using both FFPE and rapid extract method. CONCLUSION: Multiplexed interrogation for sarcoma-specific fusion transcripts using NanoString technology is a reliable approach for molecular diagnosis of pediatric sarcomas and works well with FFPE tissues. Future work will involve validating additional sarcoma fusion transcripts as well as determining the optimal workflow for diagnostic purposes.

Cardiac Findings in Fetal and Pediatric Autopsies: A 15-Year Retrospective Review.

INTRODUCTION: Congenital heart defects (CHDs) carry significant morbidity and mortality in pediatric patients. This study determined the spectrum of CHDs based on fetal and pediatric autopsies. METHODS: Autopsy reports over a 15-year period were reviewed. Postmortem findings were correlated with echocardiography records. RESULTS: From 608 autopsies, 119 cases with CHDs were identified (11% of fetal, 53% of neonatal, 18% of infant, and 4.5% of childhood autopsies). Persistent left superior vena cava was the most common individual defect. 41% of cases had extracardiac malformations. 18.5% of cases had chromosomal abnormalities. Prenatal echocardiography was available in 52 cases, showing 85% correlation with autopsy findings. Defects missed by echocardiography were generally of mild severity. CONCLUSION: Postmortem examination is important to delineate the anatomy of CHDs, and recognize extracardiac malformations for identification of possible genetic syndromes. This information can be used for parental counseling and for assessment of accuracy of pre-mortem imaging studies.

Anaplastic sarcomas of the kidney are characterized by DICER1 mutations.

Anaplastic sarcoma of the kidney is a rare tumor (≤25 reported cases) characterized by the presence of cysts, and solid areas composed of bundles of undifferentiated spindle cells, showing marked cellular anaplasia (usually accompanied by TP53 overexpression). These tumors often feature prominent areas of cartilage or chondroid material. Germline mutations in DICER1, encoding the microRNA (miRNA) processor DICER1, cause an eponymous syndrome. Recent reports suggest that anaplastic sarcoma of the kidney should be included in DICER1 syndrome as germline DICER1 mutations are associated with the occurrence of such tumors. Therefore, we sought to determine the following: (1) what proportion of anaplastic sarcoma of the kidney have DICER1 mutations; (2) whether the identified mutations affect both alleles of DICER1 (ie, are biallelic); (3) whether somatic missense mutations in the DICER1 RNase IIIb domain impact miRNA generation; and (4) whether TP53 alteration always occurs in these tumors. DICER1 mutations were evaluated by Sanger sequencing and next-generation sequencing in nine tumor/normal pairs. Impact of DICER1 mutations on miRNA generation was evaluated via an in vitro DICER1 cleavage assay. TP53 status was assessed by immunohistochemistry and next-generation sequencing. Eight of the nine cases had at least one RNase IIIb DICER1 mutation that impacted the generation of miRNAs. There were six tumors with truncating DICER1 mutations and in four of them, the mutation found in the tumor was also detected in adjacent normal tissue, and therefore was likely to be either mosaic or germline in origin. Analysis of mutation phase revealed that two of three tumors had biallelic DICER1 mutations. Six of nine anaplastic sarcomas of the kidney had aberrant TP53 immunohistochemisty with damaging TP53 mutations identified in three cases. Taken together, these data suggest that the great majority of anaplastic sarcomas of the kidney have DICER1 mutations and confirm that these tumors are part of the DICER1 syndrome.

Heterogeneity of MYCN amplification in neuroblastoma at diagnosis, treatment, relapse, and metastasis.

Amplification of the MYCN gene in neuroblastoma is associated with a poor prognosis and is considered to remain unchanged in post-treatment specimens and metastases. While heterogeneity of MYCN copy number in tumor cells has been reported, serial samples have only been studied in a limited way, and the biologic relevance of this finding is not well understood. We used in situ hybridization on paraffin sections of 102 specimens from 30 patients with MYCN-amplified neuroblastoma to determine MYCN copy number in the primary tumor, pre- and post-treatment, and in metastatic samples. Nineteen cases (63%) showed diffuse MYCN amplification in all samples tested. Nine cases (30%) showed a reduction in MYCN copy number: five cases with diffuse amplification subsequently showed focal amplification, one case with diffuse MYCN amplification showed MYCN gain after treatment, and three focally amplified cases were non-amplified in later specimens. In two cases (7%), focal amplification became diffuse in subsequent samples. Histology was not predictive of the temporal or spatial pattern of MYCN amplification for a particular tumor. If extent of amplification (focal vs. diffuse) is not considered, 26/30 (87%) of cases were consistently MYCN-amplified. However, our data suggest that MYCN status can be heterogeneous between tumor sites, during tumor progression or following treatment, challenging the notion that MYCN copy number does not change for a particular neuroblastoma. Assessing the biologic significance of MYCN heterogeneity will require larger studies of clinically annotated tumor samples, and will depend on interpreting heterogeneity in MYCN status in combination with other genetic changes. © 2016 Wiley Periodicals, Inc.

Sequencing of DICER1 in sarcomas identifies biallelic somatic DICER1 mutations in an adult-onset embryonal rhabdomyosarcoma.

BACKGROUND: Sarcomas are rare and heterogeneous cancers. We assessed the contribution of DICER1 mutations to sarcoma development. METHODS: The coding region of DICER1 was sequenced in 67 sarcomas using a custom Fluidigm Access Array. The RNase III domains were Sanger sequenced in six additional sarcomas to identify hotspot DICER1 variants. RESULTS: The median age of sarcoma diagnosis was 45.7 years (range: 3 months to 87.4 years). A recurrent embryonal rhabdomyosarcoma (ERMS) of the broad ligament, first diagnosed at age 23 years, harboured biallelic pathogenic somatic DICER1 variants (1 truncating and 1 RNase IIIb missense). We identified nine other DICER1 variants. One somatic variant (p.L1070V) identified in a pleomorphic sarcoma and one germline variant (c.2257-7A>G) may be pathogenic, but the others are considered to be benign. CONCLUSIONS: We show that deleterious DICER1 mutations underlie the genetic basis of only a small fraction of sarcomas, in particular ERMS of the urogenital tract.

C3 Glomerulopathy.

Recent advances in our understanding of the disease pathology of membranoproliferative glomerulonephritis has resulted in its re-classification as complement C3 glomerulopathy (C3G) and immune complex-mediated glomerulonephritis (IC-GN). The new consensus is based on its underlying pathomechanism, with a key pathogenetic role for the complement alternative pathway (AP), rather than on histomorphological characteristics. In C3G, loss of AP regulation leads to predominant glomerular C3 deposition, which distinguishes C3G from IC-GN with predominant immunoglobulin G staining. Electron microscopy further subdivides C3G into C3 glomerulonephritis and dense deposit disease depending on the presence and distribution pattern of electron-dense deposits within the glomerular filter. Mutations or autoantibodies affecting the function of AP activators or regulators, in particular the decay of the C3 convertase (C3 nephritic factor), have been detected in up to 80 % of C3G patients. The natural outcome of C3G is heterogeneous, but 50 % of patients progress slowly and reach end-stage renal disease within 10-15 years. The new classification not only marks significant advancement in the pathogenic understanding of this rare disease, but also opens doors towards more specific treatment with the potential for improved outcomes.

Cytokeratin-Negative Undifferentiated (Lymphoepithelial) Carcinoma of the Lacrimal Sac.

Epstein-Barr virus-associated undifferentiated (lymphoepithelial) carcinoma is a malignancy that most commonly arises in the nasopharynx but can also occur in other locations including the lacrimal sac. Generally, this tumor strongly expresses cytokeratin, making the diagnosis straightforward. In the absence of confirmatory immunohistochemistry, the diagnosis can be problematic, particularly for tumors arising in unusual locations. Only 3 cases arising in the lacrimal sac in association with Epstein-Barr virus have been reported in the English literature, and all showed typical pathologic findings. The authors report a fourth case, unique in that it showed negative immunostaining for all cytokeratins tested. The clue to the nature of the tumor came from identification of Epstein-Barr virus by in-situ hybridization and demonstration of tonofilaments by electron microscopy. This case demonstrates that a multimodal approach may be needed in the diagnosis of Epstein-Barr virus-associated carcinoma, especially when occurring in uncommon locations.