Predictive biomarkers and tumor microenvironment in female genital melanomas: a multi-institutional study of 55 cases.

Female genital melanomas are rare. At diagnosis, most affected patients have advanced disease. Surgery remains the primary treatment, and adjuvant therapy is largely ineffective. Recently, immune checkpoints and the mitogen-activated protein kinase pathway have been explored as treatment targets. However, evaluation of these biomarkers in genital melanomas is limited. We evaluated the clinicopathological features of 20 vulvar, 32 vaginal, and three cervical melanomas and assessed programmed cell death ligand 1 (PD-L1) expression, CD8 tumor-infiltrating lymphocyte density, mismatch repair proteins, VE1 immunohistochemistry, and KIT and BRAF mutations. The median age of the patients was 66 years, and median tumor sizes were 25, 30, and 20 mm for vulvar, vaginal, and cervical tumors, respectively. Mean mitotic figures were 18, 19, and 30 per mm2. Thirty-seven patients (67%) had operable tumors. After a median follow-up of 15 months, only nine patients (16%) were alive. Eight of the nine survivors did not have lymph node metastasis. Using 5% as the threshold, PD-L1 expression was observed in 55%, 50%, and 33% of vulvar, vaginal, and cervical tumors, respectively, when the Roche SP263 antibody was used and 20%, 53%, and 0%, respectively, when the Dako 28-8 antibody was used. The median CD8 tumor-infiltrating lymphocyte density was significantly higher in vulvar/vaginal than cervical melanomas and correlated with PD-L1 expression. No cases exhibited loss of mismatch repair proteins. Five cases harbored KIT mutations, three of which were hotspots. BRAF V600E mutation was not detected. Univariable analysis showed that tumor size greater than or equal to 33 mm, mitotic figures of greater than or equal to 10 per mm2, lymph node metastasis, and low CD8+ tumor-infiltrating lymphocyte density were adverse prognostic factors. Thus, patients with genital melanomas have a poor prognosis, and evaluation of multiple biomarkers is necessary to identify patients who may benefit from immunotherapy or targeted therapy.

Plasma DNA tissue mapping by genome-wide methylation sequencing for noninvasive prenatal, cancer, and transplantation assessments.

Plasma consists of DNA released from multiple tissues within the body. Using genome-wide bisulfite sequencing of plasma DNA and deconvolution of the sequencing data with reference to methylation profiles of different tissues, we developed a general approach for studying the major tissue contributors to the circulating DNA pool. We tested this method in pregnant women, patients with hepatocellular carcinoma, and subjects following bone marrow and liver transplantation. In most subjects, white blood cells were the predominant contributors to the circulating DNA pool. The placental contributions in the plasma of pregnant women correlated with the proportional contributions as revealed by fetal-specific genetic markers. The graft-derived contributions to the plasma in the transplant recipients correlated with those determined using donor-specific genetic markers. Patients with hepatocellular carcinoma showed elevated plasma DNA contributions from the liver, which correlated with measurements made using tumor-associated copy number aberrations. In hepatocellular carcinoma patients and in pregnant women exhibiting copy number aberrations in plasma, comparison of methylation deconvolution results using genomic regions with different copy number status pinpointed the tissue type responsible for the aberrations. In a pregnant woman diagnosed as having follicular lymphoma during pregnancy, methylation deconvolution indicated a grossly elevated contribution from B cells into the plasma DNA pool and localized B cells as the origin of the copy number aberrations observed in plasma. This method may serve as a powerful tool for assessing a wide range of physiological and pathological conditions based on the identification of perturbed proportional contributions of different tissues into plasma.

Applications of genetic-epigenetic tissue mapping for plasma DNA in prenatal testing, transplantation and oncology.

We developed genetic-epigenetic tissue mapping (GETMap) to determine the tissue composition of plasma DNA carrying genetic variants not present in the constitutional genome through comparing their methylation profiles with relevant tissues. We validated this approach by showing that, in pregnant women, circulating DNA carrying fetal-specific alleles was entirely placenta-derived. In lung transplant recipients, we showed that, at 72 hr after transplantation, the lung contributed only a median of 17% to the plasma DNA carrying donor-specific alleles, and hematopoietic cells contributed a median of 78%. In hepatocellular cancer patients, the liver was identified as the predominant source of plasma DNA carrying tumor-specific mutations. In a pregnant woman with lymphoma, plasma DNA molecules carrying cancer mutations and fetal-specific alleles were accurately shown to be derived from the lymphocytes and placenta, respectively. Analysis of tissue origin for plasma DNA carrying genetic variants is potentially useful for noninvasive prenatal testing, transplantation monitoring, and cancer screening.

Axillary Nodal Metastasis with Papillary Morphology: An Uncommon Origin.

BACKGROUND: Metastases to axillary lymph nodes are commonly and readily confirmed by fine needle aspiration cytology (FNAC). Most likely, these arise from breast primaries. However, the diagnosis can become complicated when unusual cytomorphology is encountered. CASE: We report a 60-year-old woman presenting with bilateral axillary lymphadenopathies but without breast lesions. History showed increasing CA-125 levels. FNAC yielded carcinoma cells showing prominent papillary pattern, being composed of mild to moderately differentiated malignant cells, with focal abortive glandular formation and squamous metaplasia. IHC stains were done and the tumor cells were PAX-8 positive, but GATA-3 and GCDFP-15 negative. Coupled with the clinical history, a diagnosis of metastatic endometrioid adenocarcinoma was made. CONCLUSION: Nodal metastases with papillary cytomorphology can rarely arise from nonbreast primaries. The presence of papillary pattern, particularly in the absence of a clinically detectable breast lesion, should raise the possibility of a metastasis. Correlation with patient history, imaging findings and judicious use of IHC studies are crucial for a correct diagnosis.

Cutaneous hemangioma in a giant panda (Ailuropoda melanoleuca).

An adult male giant panda (Ailuropoda melanoleuca) was presented with a cutaneous mass ventral to the eye. The animal was anesthetized and the mass was surgically excised. Histopathologic examination determined that the mass was a benign cavernous hemangioma, the first reported case in a giant panda.

Necrotizing enterocolitis complicated with perforation in extremely low birth-weight premature infants.

This study determined the incidence, clinical characteristics, treatment and outcome in extremely low birth-weight (ELBW) premature infants with perforated necrotizing enterocolitis (NEC). We retrospectively reviewed the medical records of ELBW (birth weight <1000 g ) premature infants with perforated NEC diagnosed and managed at National Taiwan University Hospital (NTUH) from January 1993 through December 2000. A total of 8 ELBW premature infants with perforated NEC were collected. The incidence of perforated NEC in ELBW premature infants was 5.1% (8 out of 158). The average age at onset of perforated NEC was 26 days. The most common clinical features were abdominal distention, decreased bowel sound and poor activity level. Dilated and fixed bowel loops, bowel wall thickening and ascites with stool-like substance drainage out from penrose drain tube were the predominant signs at the time of diagnosis of perforated NEC. Thrombocytopenia, elevated C-reactive protein and anemia were the major laboratory findings. All infants received a primary penrose drain in the acute stage of disease. The overall survival rate was 37.5% (3 out of 8). Death occurred due to nosocomial infection with sepsis in 3 patients and due to perforated NEC in 2 patients. Two of the three surviving patients started enteral feeding 19 and 41 days after the diagnosis of perforated NEC and tolerated oral feedings well; the third patient still required total parenteral nutrition two years after diagnosis. Although the clinical characteristics and radiographic findings of perforated NEC in ELBW premature infants were variable, brown color ascites with stool-like substance may be considered a significant sign of perforated NEC despite the absence of free air on radiography at the early stage of disease. Close observation of clinical symptoms and signs, more aggressive surgical intervention and prevention of the following nosocomial infection may have the opportunity to reduce the mortality due to perforated NEC.

Aberrant large tumor suppressor 2 (LATS2) gene expression correlates with EGFR mutation and survival in lung adenocarcinomas.

BACKGROUND: Large tumor suppressor 2 (LATS2) gene is a putative tumor suppressor gene with potential roles in regulation of cell proliferation and apoptosis in lung cancer. The aim of this study is to explore the association of aberrant LATS2 expression with EGFR mutation and survival in lung adenocarcinoma (AD), and the effects of LATS2 silencing in both lung AD cell lines. METHODS: LATS2 mRNA and protein expression in resected lung AD were correlated with demographic characteristics, EGFR mutation and survival. LATS2-specific siRNA was transfected into four EGFR wild-type (WT) and three EGFR mutant AD cell lines and the changes in LATS2 expression and relevant signaling molecules before and after LATS2 knockdown were assayed. RESULTS: Fifty resected lung AD were included (M:F=23:27, smokers:non-smokers=19:31, EGFR mutant:wild-type=21:29) with LATS2 mRNA levels showed no significant difference between gender, age, smoking and pathological stages while LATS2 immunohistochemical staining on an independent set of 79 lung AD showed similar trend. LATS2 mRNA level was found to be a significant independent predictor for survival status (disease-free survival RR=0.217; p=0.003; Overall survival RR=0.238; p=0.036). siRNA-mediated suppression of LATS2 expression resulted in augmentation of ERK phosphorylation in EGFR wild-type AD cell lines with high basal LATS2 expression, discriminatory modulation of Akt signaling between EGFR wild-type and mutant cells, and induction of p53 accumulation in AD cell lines with low baseline p53 levels. CONCLUSIONS: LATS2 expression level is predictive of survival in patients with resected lung AD. LATS2 may modulate and contribute to tumor growth via different signaling pathways in EGFR mutant and wild-type tumors.

Evaluation of 2 real-time PCR assays for in vitro diagnostic use in the rapid and multiplex detection of EGFR gene mutations in NSCLC.

Activating mutations of the epidermal growth factor receptor (EGFR) gene in non-small cell lung cancer predict for a favorable clinical response to tyrosine kinase inhibitor therapy. Although Sanger sequencing is a conventional method to detect EGFR gene mutations, multiplex real-time allele-specific polymerase chain reaction (PCR) systems are increasingly used in the routine molecular diagnostic setting. We aim to evaluate 2 proprietary real-time PCR assays (cobas and therascreen) against Sanger sequencing in the detection of EGFR gene mutations. The overall concordance rate between cobas and therascreen assays with Sanger sequencing was 89% and 88%, respectively, and increased to 96% and 98%, respectively, if the mutations not covered were excluded. The cobas assay showed a superior coverage of exon 20 mutations, but L861Q was not targeted. The nature of specimen, DNA integrity, and tumor cell content are factors that affect the assay performance. DNA extracted from cell block and clot of pleural fluid gave rise to 1 invalid call and 1 false-negative result by the cobas assay and 1 missed T790M mutation and 1 false-negative result by the therascreen assay. Both assays are around 5 times more expensive compared with Sanger sequencing in terms of reagent cost. We conclude that both assays prove to be a rapid, simple, and validated method in detecting the most common and clinically significant EGFR gene mutations in non-small cell lung cancer. Although less convenient compared with real-time PCR assays, Sanger sequencing is cheaper in terms of reagent cost and allows the detection of rare or novel EGFR gene mutations.

Dual-tracer PET/CT in renal angiomyolipoma and subtypes of renal cell carcinoma.

UNLABELLED: We studied the metabolic characteristics of RCC subtypes and angiomyolipoma with 18F-FDG and 11C-acetate PET/CT. METHODS: Fifty-eight patients with both baseline CT and dual-tracer PET/CT were recruited: 10 angiomyolipoma (16 lesions) and 48 RCC (50 lesions). Each lesion was assessed for SUVmax ratio (lesion-to-normal kidney) on 11C-acetate/18F-FDG PET and attenuation density on CT. Receiver operating characteristic (ROC) curve was analyzed to define the threshold of 11C-acetate SUVmax ratio for differentiating angiomyolipoma from RCC. Thirty-nine RCC patients were selected for 3-year disease-free survival analysis. RESULTS: All angiomyolipoma showed negative 18F-FDG but markedly increased 11C-acetate metabolism, significantly higher than RCC (11C-acetate SUVmax ratio = 4.11 ± 0.53 vs 2.00 ± 0.71; P < 0.05). 11C-acetate SUVmax ratio = 3.71 could differentiate angiomyolipoma including "fat-poor angiomyolipoma" (n = 10) from RCC with sensitivity of 93.8% (15/16) and specificity of 98.0% (49/50). Different RCC subtypes/grades (25 low- and 11 high-grade clear cell [CC], 7 chromophobe, 4 papillary, and 1 collecting duct) were found to have different dual-tracer metabolic pattern (P < 0.05), with overall RCC detection sensitivity of 90% (45/50). All chromophobe RCC were avid only for C-acetate but not 18F-FDG, whereas papillary RCC were primarily the opposite. RCC-CC showed variable dual-tracer uptake: high-grade more avid for F-FDG, low-grade more for 11C-acetate. Four RCC cases negative by dual-tracers were of low-grade RCC-CC. "Primary RCC being 18F-FDG-avid" was the only independent predictor of RCC recurrence in 3 years (P < 0.05), with a median disease-free survival of 22 months. CONCLUSION: 11C-acetate PET/CT helps in differentiating "fat-poor angiomyolipoma" from RCC. Dual-tracer PET/CT has value in diagnosis of RCC subtypes and predicting survival.

Type II enteropathy-associated T-cell lymphoma: a distinct aggressive lymphoma with frequent γδ T-cell receptor expression.

Enteropathy-associated T-cell lymphoma (EATL), an uncommon lymphoma of intestinal intraepithelial T lymphocytes, occurs with a higher frequency in northern Europe due to association with celiac disease. Data on the occurrence of EATL in the Asian population, among whom celiac disease is very rare, are conflicting. This study aimed to characterize EATL encountered in the Chinese population in Hong Kong. Eighteen cases were identified, all fulfilling the criteria of type II rather than classical EATL. The patients, including 13 men and 5 women, had a median age of 62 years. Most presented with small bowel perforation, and there was no history of malabsorption. The clinical course was aggressive, with 14 of 16 patients dying of progressive disease or complications, usually within 1 year. The histologic features were practically identical in all cases. The central zone of the tumor showed ulceration with or without perforation and was characterized by monotonous transmural infiltration of the bowel by small-sized or medium-sized lymphoma cells with few admixed inflammatory cells and no coagulative necrosis. The peripheral zone featured lateral spread of lymphoma cells in the mucosa, accompanied by variable involvement of the submucosa and muscularis. In all cases, there was an intraepithelial lymphocytosis zone contiguous or discontinuous with the peripheral zone, which was characterized by infiltration of the intestinal epithelium by nonatypical small lymphocytes, and not accompanied by other histologic changes of enteropathy. The most common phenotype of the lymphoma cells was CD3+, CD5-, CD4-, CD8+, CD56+, TIA1+, CD30-, and Epstein-Barr virus, and 2 cases showed aberrant expression of CD20. A remarkable finding was that 14 (78%) cases expressed γδ T-cell receptor, and only 6 (33%) expressed αß T-cell receptor (with 3 cases coexpressing both T-cell receptors and 1 case expressing neither). The immunophenotype of the intraepithelial lymphocytes was either discordant (particularly with respect to CD8 and CD56 expressions) or concordant with the lymphoma cells of the corresponding cases. Thus, this study shows that EATL occurring in the Chinese population is exclusively of type II. In contrast to several studies, intraepithelial lymphocytosis can be consistently demonstrated and this component seems to represent a precursor lesion of EATL rather than a manifestation of celiac disease. In view of the differences in epidemiology and clinicopathologic features, we believe it is justified to separate out type II EATL from the EATL category as a distinct form of lymphoma, for which we propose the designation "monomorphic intestinal T-cell lymphoma."

Idiopathic cervical fibrosis--a new member of IgG4-related sclerosing diseases: report of 4 cases, 1 complicated by composite lymphoma.

Idiopathic cervical fibrosis is a rare tumefactive inflammatory-sclerosing lesion involving the soft tissues of the head and neck, and a proportion of patients also have synchronous or metachronous inflammatory fibrosclerosing lesions in other anatomic sites. The latter finding suggests that this entity may represent a member of IgG4-related sclerosing diseases. We report 4 cases to support this postulation. The patients were male adults aged 42 to 89 years, who presented with an infiltrative, firm cervical mass. Two patients also had IgG4-related chronic sclerosing sialadenitis of submandibular gland and lymphadenopathy. Histologically, the cervical soft tissue lesions had ill-defined borders, consisting of coalescent nodular lymphoid aggregates accompanied by a sclerotic stroma. Nerve infiltration, skeletal muscle invasion, and phlebitis were present. There was a significant increase in IgG4 plasma cells (87 to 327 per high-power field, with IgG4/IgG ratio of 63% to 98%). In the soft tissue lesion of 1 patient, there were expansile foci comprising dense sheets of plasma cells and small lymphoid cells that exhibited κ light chain restriction and clonal immunoglobulin gene rearrangement, consistent with supervening extranodal marginal zone lymphoma. The adjacent lymph node from the same patient showed Epstein-Barr virus (EBV)-positive classical Hodgkin lymphoma with typical morphology and immunophenotype (CD30, CD15, PAX5). Thus lymphoma can supervene in the chronic inflammatory background similar to that recently documented for IgG4-related sclerosing disease of the ocular adnexa.

Spontaneous atraumatic rupture of a normal-sized spleen due to AL amyloid angiopathy.

We report an unusual cause of hemoperitoneum in an AL amyloid patient on peritoneal dialysis due to spontaneous rupture of a normal-sized spleen not related to any trauma. The rupture was not due to amyloid deposition within the spleen pulp but rather due to amyloid angiopathy causing hemorrhage within the spleen and capsular tear.

Ocular adnexal lymphoma associated with IgG4+ chronic sclerosing dacryoadenitis: a previously undescribed complication of IgG4-related sclerosing disease.

IgG4-related sclerosing disease is a recently recognized inflammatory lesion frequently involving pancreas, submandibular gland, lacrimal gland, and lymph node. We report 3 cases of ocular adnexal lymphoma arising in IgG4-related chronic sclerosing dacryoadenitis, a phenomenon that has not been previously reported. The patients presented with bilateral or unilateral ocular adnexal mass usually present for many years. One patient also had asymptomatic diffuse lymphadenopathy. Two patients had biopsy-proven IgG4-related chronic sclerosing dacryoadenitis before the current presentation, and 1 had systemic involvement by IgG4-related sclerosing disease as evidenced by increased IgG4+ cells in a prior nasopharyngeal biopsy. Two cases showed features of extranodal marginal zone lymphoma of mucosa-associated lymphoid-tissue type (1 with large cell transformation) and 1 follicular lymphoma. Thus, the lymphoid hyperplasia of IgG4-related sclerosing disease can provide a substrate for the emergence of lymphoma. In addition, we report 3 cases of ocular adnexal extranodal marginal zone B-cell lymphoma that show sclerosing inflammation in the background and numerous IgG4+ monotypic plasma cells. In the absence of prior biopsies or information on serum IgG4 titer, it is unclear whether these cases represent lymphoma complicating IgG4-related sclerosing disease or de novo lymphoma. Nonetheless, these cases are distinctive in that the neoplastic cells express IgG4 (light chain restricted), whereas unselected cases of ocular adnexal lymphomas do not show IgG4 expression.

Deregulation of E-cadherin-catenin complex in precancerous lesions of gastric adenocarcinoma.

BACKGROUND AND AIM: Decrease in expression of the E-cadherin-catenin complex is an important element in gastric carcinogenesis. However, the expression of the complex in gastric precancerous lesions has not been well studied. The present study aimed to examine the serial change in expression of E-cadherin-catenin complex in the precancerous lesions of gastric cancer patients. METHODS: Gastrectomy specimens of 40 patients with gastric cancer were retrieved. Areas with chronic gastritis, atrophic gastritis, intestinal metaplasia and adenocarcinoma were identified and immunostained for alpha-catenin, beta-catenin and E-cadherin. The results were scored semiquantitatively by two independent pathologists using a validated scoring system. RESULTS: A significant decrease in score was observed in 5% (1/22) of alpha-catenin, 0% (0/22) of beta-catenin and 9% (2/22) of E-cadherin in chronic atrophic gastritis patients, and in 28% (5/18) of alpha-catenin, 67% (10/15) of beta-catenin and 57% (8/14) of E-cadherin in intestinal metaplasia patients. The scoring of alpha-catenin, beta-catenin and E-cadherin correlated with each other. Forty-three percent of patients had concordant changes of scores along the gastritis-adenocarcinoma sequence. There was no association between Helicobacter pylori status and E-cadherin-catenin complex expression. CONCLUSION: Deregulation of the E-cadherin-catenin complex was observed in the majority of precancerous lesions in patients with gastric adenocarcinoma, which has potential diagnostic and therapeutic implications.