Expression of B7-H3 and B7-H4 in Gastric Gastrointestinal Stromal Tumors.

Gastric gastrointestinal stromal tumors (GISTs) are mesenchymal neoplasms with variable behavior characterized by differentiation toward the interstitial cells of Cajal occurring anywhere in the gastrointestinal stromal tract. The management of GIST was revolutionized by the introduction of imatinib, a KIT inhibitor, which has become the standard first-line treatment for metastatic GIST. However, despite a clinical benefit rate of 80%, the majority of patients with GIST experience disease progression after 2 to 3 years of imatinib therapy. This shows the need for novel treatment approaches for imatinib refractory GISTs. The checkpoint proteins B7-H3 and B7-H4 inhibit the activation and function of T cells by potently suppressing the proliferation, cytokine production, and cytotoxicity of activated T cells, which is a mechanism for immune escape. This study aims to clarify B7-H3 and B7-H4 expression in gastric GISTs using immunohistochemistry and reverse transcription polymerase chain reaction (RT-PCR). We confirmed B7-H3 expression (H-score ≥50 points) in 92% and B7-H4 expression in 0% of GIST samples. We examined B7-H3 mRNA expression in 3 representative GIST samples, each having their respective immunostained areas detected by RT-PCR. B7-H3 is expressed at a particularly high rate in GISTs. This suggests that B7-H3 might operate as part of an immune checkpoint in GISTs.

Resolution of Breast Cancer in a Patient With Thyroid Stimulating Hormone-Secreting Pituitary Neuroendocrine Tumor With the Combination of Chemotherapy and Lanreotide.

Thyroid stimulating hormone-secreting pituitary neuroendocrine tumor (TSH-PitNET) is a rare disease in which pituitary adenomas secrete excessive amounts of TSH, and TSH is not suppressed despite high blood levels of thyroid hormone. Somatostatin analogs (SSAs) like lanreotide are used to control TSH secretion and manage symptoms in cases where surgery is not fully effective or feasible. The treatment of choice for human epidermal growth factor 2 receptor (HER2)-positive metastatic breast cancer is generally chemotherapy and anti-HER2 therapy. A 52-year-old woman was diagnosed with Graves' disease 26 years ago and stopped going to the hospital after several years of treatment with thiamazole. She had a right breast mass two years prior and visited the Department of Breast and Endocrine Surgery in our hospital one year prior, where she was diagnosed with T3N3M1, stage 4 breast cancer with a mass 52 mm in diameter in the right breast and metastasis in the 12th thoracic vertebra. Breast cancer receptor status was negative for the estrogen receptor, negative for the progesterone receptor, and positive for HER2. She was also found to have an enlarged thyroid gland, palpitations, inappropriate TSH secretion, and a 6 mm nodule on the pituitary gland, which was diagnosed as a TSH-PitNET. She was treated for breast cancer with trastuzumab deruxtecan therapy and for TSH-PitNET with lanreotide. One month after starting lanreotide, pituitary, and thyroid function improved to normal, and four months later, the breast mass was significantly reduced to 16 mm in diameter and a mastectomy was performed. The size of the pituitary adenoma remained unchanged during observation. Remarkably, the mastectomy specimen was free of cancer cells and showed a pathologically complete response. Needle biopsy specimens at the time of breast cancer diagnosis were positive for somatostatin receptor 2 (SSTR2) and insulin-like growth factor 1 receptor (IGF-1R) immunostaining. However, both were negative in the mastectomy specimen. Recently, SSTR2 and IGF-1R were reported to be expressed in breast cancer, and several clinical trials of SSAs for breast cancer have been conducted. SSAs are effective in improving pituitary and thyroid functions against TSH-PiTNET, and in combination with chemotherapy, they may have synergistic antitumor effects in patients with SSTR2-positive breast cancer.

GATA4 is diagnostically useful for distinguishing primary ovarian mucinous carcinomas from metastatic colorectal adenocarcinomas to the ovary.

AIM: Determining the primary origin of an ovarian mucin-producing carcinoma can be challenging at times because some metastases of primary colorectal origin may exhibit gross, microscopic, and/or immunohistochemical features that overlap with those of primary ovarian mucinous carcinomas (OMCs). We hypothesized that GATA binding protein 4 (GATA4) might be a novel, useful marker for differentiating primary OMCs from metastatic colorectal adenocarcinomas to the ovary. METHODOLOGY: For comparison with the usefulness of other markers (special AT-rich sequence-binding protein 2 (SATB2) and caudal type homeobox 2 (CDX2)), we elucidated the expression profiles of GATA4 in OMCs, colorectal non-mucinous adenocarcinomas (CNMACs), and colorectal mucinous adenocarcinomas (CMACs) using immunohistochemistry. RESULTS: We confirmed GATA4 expression (H-score ≥50 points) in 93%, SATB2 in 0%, and CDX2 in 64% of 14 OMCs. GATA4 was expressed in 13%, SATB2 in 90%, and CDX2 in 93% of 30 CNMACs. GATA4 was expressed in 20%, SATB2 in 73%, and CDX2 in 100% of 30 CMACs. CONCLUSION: The expression of GATA4 in a mucus-producing ovarian tumor strongly supports it being a primary OMC rather than a metastatic colorectal carcinoma: GATA4 expression indicates OMC and SATB2 expression indicates colorectal adenocarcinoma. However, three cases of colorectal adenocarcinoma were GATA4-positive and SATB2-negative, so the GATA4/SATB2 marker combination is not absolute for determining the primary site. Further research for more markers is necessary to find the ideal combination.

Case report: Atypical case of autoimmune glial fibrillary acidic protein astrocytopathy following COVID-19 vaccination refractory to immunosuppressive treatments.

A 54-year-old Japanese man presented with headache and fever the day after SARS-CoV-2 vaccination. He became deeply unconscious within a week. Brain MRI showed periventricular linear enhancements and a few spotty lesions in the cerebral white matter. Cerebrospinal fluid (CSF) testing showed mild pleocytosis. He was treated with intravenous methylprednisolone and plasma exchange. However, the white matter lesions enlarged to involve the brainstem and cerebellum, and long cord spinal lesions appeared. Anti-glial fibrillary acidic protein (GFAP) antibody was positive in the CSF and serum, and he was therefore diagnosed as autoimmune GFAP-astrocytopathy (GFAP-A). In addition, high-dose immunoglobulin therapy was administered twice, but his symptoms did not improve; the white matter lesions enlarged further, and modified Rankin Scale score increased to 5. A brain biopsy specimen showed infiltration of macrophages and CD4 + lymphocytes together with neuron and oligodendrocytic injuries and glial scar. Although GFAP-A generally responds well to steroids, the present case developed GFAP-A following SARS-CoV-2 vaccination, with refractory to intensive immunosuppressive therapy and atypical pathologic findings of infiltration of CD4 + lymphocytes and demyelination.

Expression of TGF-ß1 in Gastrointestinal Stromal Tumor (GIST) and the Occurrence of Frequent Desmoplasia.

Gastrointestinal stromal tumors (GISTs) are mesenchymal neoplasms with variable behavior characterized by differentiation toward the interstitial cells of Cajal occurring anywhere in the gastrointestinal stromal tract. Frequently, GISTs have fibrous stroma within tumor cell proliferation areas, which is unlike other types of malignant tumors. If this desmoplasia is active, there is a possibility that some sort of transmitter exists between GIST cells and cells related to fibrosis in the tumor cell proliferation areas. Transforming growth factor (TGF)-ß isoforms, particularly TGF-ß1, are critical for fibrosis pathogenesis. TGF-ß1 regulation of myofibroblasts and fibroblasts during fibrosis is well described. The induced fibroblast activation resulting in myofibroblast differentiation has been reported as an important source of collagen, glycoproteins, proteoglycans, and matrix metallopeptidases in wound healing and fibrosis. However, there are a few reports on the relationship between TGF-ß1 and GISTs. This study aims to clarify TGF-ß1 expression in 30 gastric GISTs using immunohistochemistry and reverse transcription polymerase chain reaction (RT-PCR). For comparison, we also enrolled 30 samples of gastric tubular adenocarcinoma (GTAC). We confirmed TGF-ß1 expression (H-score ≥50 points) in 57% of GIST and 13% of GTAC samples, a significant difference between the 2 tumor types ( P =0.001). We examined the TGF-ß1 mRNA expression of 3 representative GIST samples, each having their respective immunostained areas detected by RT-PCR. Finding TGF-ß1 expression may indicate that this cytokine plays a part in the formation of desmoplasia within GIST cell proliferative areas.

Expression of CXCL12 in esophageal high grade dysplasia characterized pathologically by lymphocyte accumulation directly under the lesion.

Squamous dysplasia of the esophagus is an unequivocal neoplastic alteration of the esophageal squamous epithelium without invasion. Esophageal high grade dysplasia (EHGD) is characterized by >50% epithelial involvement or severe cytological atypia. Frequently, lymphocytes accumulate below EHGD lesions even though there is no invasion. If this lymphocytic accumulation is active, a transmitter should exist between the EHGD cells and the lymphocytes. C-X-C motif chemokine ligand (CXCL) 12, CXCL10 and C-C motif chemokine ligand 18 (CCL18) are all lymphocyte chemoattractants in vivo, but there are no reports on the relationship between these chemokines and EHGDs. In this study, we investigated these chemokines and C-X-C motif chemokine receptor 4 (CXCR4) (receptor for CXCL12) in 30 EHGDs using immunohistochemistry and reverse transcription polymerase chain reaction (RT-PCR). For comparison, we enrolled 30 samples of normal esophageal squamous epithelium (NESE). We confirmed CXCL12 expression (H-score≥50 points) in 70% of EHGD and 0% of NESE samples, CXCL10 expression in 3% of EHGD and 3% of NESE samples, CCL18 expression in 3% of EHGD and 0% of NESE samples, and CXCR4 expression in 53% of EHGD and 0% of NESE samples by immunohistochemistry. EHGD and NESE cases were significantly different in their expressions between the tissue types (CXCL12, p<0.001; CXCR4, p<0.001). We examined CXCL12 and CXCR4 mRNA expressions of 3 representative EHGD samples, each having their respective immunostained areas detected by RT-PCR. Finding CXCL12 expression may indicate that this chemokine plays a part in the lymphocyte accumulation that occurs directly under EHGDs.

Expressions of CXCL12, CXCL10 and CCL18 in Warthin tumors characterized pathologically by having a lymphoid stroma with germinal centers.

The Warthin tumor is a benign neoplasm of the salivary glands, histologically, the tumor has an oncocytic epithelial component forming uniform rows of cells surrounded by cystic spaces associated with a lymphoid stroma often showing the presence of germinal centers. The lymphoid stroma is a representative microscopic finding. If this lymphocytic accumulation is active, some sort of transmitter should exist between the Warthin tumor cells and lymphocytes. C-X-C motif chemokine ligand (CXCL12) 12, CXCL10 and C-C motif chemokine ligand 18 (CCL18) are a chemoattractant for lymphocytes in vivo. There is no report on the relationship between these chemokines and Warthin tumors. In this study, we investigated these chemokines expressions in 20 Warthin tumors using immunohistochemistry and reverse transcription polymerase chain reaction (RT-PCR). For comparison, we also enrolled samples of pleomorphic adenoma, which is another benign salivary gland tumor type without prominent lymphocytic infiltration. All Warthin tumors were immunopositive for CXCL12 and CXCL10, and these reactivities were diffuse. Meanwhile, the majority of pleomorphic adenomas were immunonegative for CXCL12 (95%), CXCL10 (80%) and CCL18 (85%). Warthin tumor and pleomorphic adenoma cases were significantly different in these immunostaining expressions (CXCL12, p<0.001; CXCL10, p<0.001; CCL18, p=0.024). We examined CXCL12, CXCL10 and CCL18 mRNA expressions of 3 representative Warthin tumor samples, each having these chemokines immunopositive areas detected by RT-PCR. Finding CXCL12 and CXCL10 expressions indicate that these chemokines may play a part in the formation of a lymphoid stroma within Warthin tumors. In regards to this phenomenon, the participation of CCL18 might be restrictive compared to CXCL12 and CXCL10.

Progression of Papillary Thyroid Carcinoma to Anaplastic Carcinoma in Metastatic Lymph Nodes: Solid/Insular Growth and Hobnail Cell Change in Lymph Nodes Are Predictors of Subsequent Anaplastic Transformation.

Most anaplastic thyroid carcinomas (ATCs) arise from papillary thyroid carcinoma (PTC). This process is also called anaplastic transformation, and the morphological harbingers of this phenomenon in nodal recurrence have not been assessed systematically. For this reason, the current study focused on features of 10 PTCs with regional lymph node recurrence that was accompanied with disease progression due to anaplastic transformation in at least one of the nodal recurrences. The findings of additional 19 PTCs which recurred without anaplastic transformation after ≥ 10 years of follow-up served as the control group. There were no clinicopathological differences between the two groups at initial surgery including age, gender, tumor size, lymph node metastasis, distant metastasis, extrathyroidal extension, histologic subtype, and treatment. The median time from the initial thyroid surgery to anaplastic transformation in the nodal recurrence was 106 months (range 6 to 437 months). Mutational analyses showed recurrent PTCs with anaplastic transformation had a high prevalence of BRAFV600E mutation (8/9) and TERT promoter mutation (9/9), both of which were detected in primary tumors. PIK3CAH1047R mutation was detected in one case. No case had RAS mutation. Nineteen recurrent PTCs without anaplastic transformation harbored BRAFV600E mutation and seventeen of these had TERT promoter mutation. Unlike primary tumors with subsequent nodal anaplastic transformation, TERT promoter mutation was only present in the metastatic nodal recurrence from 4 patients without transformation. No patients had neither high-grade features (necrosis and increased mitotic activity) nor solid/insular growth or hobnail cell features in their primary tumors. In the group of patients with transformation, 3 had solid/insular growth in the lymph node metastasis at the time of primary tumor resection (one displaying nuclear features of PTC and solid growth with increased mitotic activity, one with insular component consistent with poorly differentiated carcinoma component, and one displaying nuclear features of PTC and solid growth), and additional 2 patients had solid/insular growth with no high-grade features or poorly differentiated carcinoma component at the time of subsequent nodal recurrence prior to anaplastic transformation. Hobnail cell features were exclusively seen in subsequent metastatic lymph nodes prior to anaplastic transformation. The control group lacked solid/insular growth and hobnail cell features in the metastatic nodal disease. Aberrant p53 expression and loss of TTF-1 featured tumor components with anaplastic transformation. This series identified a subset of recurrent PTCs with TERT promoter mutation was prone to undergo anaplastic transformation, and that solid/insular growth and hobnail cell features were morphological predictors of anaplastic transformation in the nodal recurrence.

Nodal EBV-positive polymorphic B cell lymphoproliferative disorder with plasma cell differentiation: clinicopathological analysis of five cases.

Plasma cell differentiation (PCD) is frequently observed in some entities of non-Hodgkin B cell lymphoma, including both low-grade and high-grade lymphomas. However, except for plasmablastic lymphoma and primary effusion lymphoma, EBV+ B cell lymphoproliferative disorder (LPD) with PCD has not been well addressed due to its rarity. We clinicopathologically examined five cases of nodal EBV+ polymorphic B cell LPD with PCD (PBLPD-PCD) initially diagnosed as polymorphic EBV+ diffuse large B cell lymphoma, not otherwise specified (DLBCL-NOS) with PCD (n = 3) and methotrexate-associated B cell LPD (MTX-associated B-LPD) (n = 2). One case had a concomitant brain lesion which was clinically diagnosed as EBV-related encephalitis. This patient received therapy with vidarabine, and both the brain lesion and the nodal EBV+ PBLPD-PCD lesions disappeared. Another case was characterized by Mott cell differentiation. This case was the first reported case of EBV+ B cell lymphoma or LPD with Mott cell differentiation. The two cases of MTX-associated B cell LPD which arose in patients with rheumatoid arthritis spontaneously regressed after MTX cessation. TCRγ and IGH PCR analysis was performed in four cases. Two cases had TCRγ rearrangements, but no IGH rearrangements. The other two cases had no rearrangements in these genes. We concluded that nodal EBV+ PBLPD-PCD is rare, with heterogeneous characteristics. PCR analysis revealed that nodal EBV+ PBLPD-PCD may have only TCR clonality and no IGH clonality. Considering the partial or complete loss of CD20 expression on the tumor cells, this result may be confusing for accurate diagnosis of EBV+ PBLPD-PCD, and pathologists need to be aware of this phenomenon to avoid misdiagnosis.

Expressions of IL-8 and CXCL5 in uterine endometrioid carcinomas which have frequent neutrophil infiltration and comparison to colorectal adenocarcinoma.

In endometrioid carcinomas (ECs) of the uterine corpus, neutrophil accumulation within the carcinoma cell clusters is a representative microscopic finding. Because this accumulation is active, some sort of transmitter ought to exist between the EC cells and neutrophils. Interleukin-8 (IL-8) and C-X-C motif chemokine ligand 5 (CXCL5) is a cytokine that attracts neutrophils in vivo. In this study, we investigated IL-8, CXCL5 and C-X-C motif chemokine receptor 2 (CXCR2) (their chemokine receptor) expressions in ECs by immunohistochemistry and reverse transcription polymerase chain reaction (RT-PCR). There are few reports on the relationship between these chemokines and ECs. For comparison, we enrolled samples of colorectal adenocarcinoma (CRAC), it is another representative tumor with neutrophil infiltration. We analyzed 30 ECs and 30 CRACs. We confirmed IL-8 expression (H-score ≥50 points) in 40% of EC and 7% of CRAC samples; CXCL5 expression in 7% of EC and 10% of CRAC samples; CXCR2 expression in 83% of EC and 53% of CRAC samples by immunohistochemistry. We examined each mRNA (IL-8 and CXCL5) expression of 3 representative EC and 3 CRAC samples. Finding IL-8 expression might indicate that this cytokine is important for the process of neutrophil accumulation, particularly within ECs. The participation of CXCL5 regarding neutrophil accumulation within their carcinoma cell clusters might be restrictive compared to IL-8.

Identification of Recurrent TERT Promoter Mutations in Intrathyroid Thymic Carcinomas.

Intrathyroid thymic carcinoma (ITTC) is a rare malignant neoplasm considered to be a eutopic thymic carcinoma (TC) arising ectopically in the thyroid. Histopathologically, ITTC resembles squamous cell carcinoma of the thymus with positive TC markers such as CD5 and c-KIT. Despite these similar histological findings, ITTC is clinically less aggressive than TC. In this study, we compared clinical, histological, and genetic characteristics of ITTCs and TCs. We collected 9 ITTCs and 8 TCs with their clinicopathological profiles. Immunohistochemistry for CD5, p63, CD117/c-KIT, Ki-67, p53, TTF-1, thyroglobulin, PAX8, EGFR, and PD-L1/CD274 plus in situ hybridization for EBER was performed. We further investigated mutation status of KIT, EGFR, BRAF, and TERT promoter using Sanger sequencing. In our study, ITTCs affected significantly younger patients than TCs. After a mean follow-up of 86 months, all patients with ITTC were alive, while two patients with TC had died. Immunohistochemistry showed ITTCs and TCs had a similar immunophenotype except for EGFR and p53. Genetic analysis did not identify KIT or BRAF mutations in any ITTCs or TCs. EGFR mutations were positive in 11% (1/9) of ITTCs and 25% (2/8) of TCs. Notably, TERT promoter C228T mutation was identified in 22% (2/9) of ITTCs but none of the TCs. There were no significant differences in age, tumor size, or sex between TERT-mutated and TERT-wild-type ITTCs. Collectively, ITTC and TC have similar histopathologic and immunophenotypic features but different clinical outcomes. Recurrent TERT promoter mutation may be a key event related to cancer progression in ITTCs and warrants further investigation.

IgG4-related disease of the paratestis with the scrotal fluid: A case report.

IgG4-related disease (IgG4-RD) can affect various organs, and the pancreas and salivary gland are representative examples. We report a rare case of IgG4-RD of the paratestis. A 74-year-old man presented with left scrotal swelling. Scrotopuncture drainage and cytology confirmed a clear, yellow retention liquid (130 mL) with many small, similar lymphocytes and a few plasmacytes. Many lymphoid cells were immunopositive for CD3 on a cell block section, indicating that a predominant type of lymphoid cells was T cell. There were also some CD20 immunopositive cells and a few IgG4 immunopositive cells. Two months later the left scrotal swelling had returned, and he underwent radical inguinal orchiectomy. Microscopically, there was considerable lymphoplasmacytic inflammatory infiltration, fibrosis and abundant IgG4 immunopositive cells in the paratesticular region. The histopathologic and immunohistochemistry findings were consistent with IgG4-RD. However, the abundant T cells in the scrotal fluid complicated the cytological diagnosis in our case.

SMARCB1/INI1 Is Diagnostically Useful in Distinguishing α-Fetoprotein-producing Gastric Carcinoma from Hepatocellular Carcinoma.

BACKGROUND/AIM: Switch/sucrose non-fermentable (SWI/SNF)-related, matrix-associated, actin-dependent regulator of chromatin subfamily B member 1 (SMARCB1), also named integrase interactor 1, is one of the core subunit proteins in the SWI/SNF ATP-dependent chromatin remodeling complex encoded at chromosomal position 22q11.2. Complete loss of SMARCB1 expression has been reported in various malignant tumors. Immunohistochemistry has demonstrated that SMARCB1 mutation/inactivation correlates well with loss of nuclear SMARCB1 expression. This study investigated SMARCB1 expression in hepatocellular carcinomas (HCCs) and α-fetoprotein (AFP)-producing gastric carcinomas by immunohistochemistry. For comparison, SMARCB1 immunostaining in intrahepatic cholangiocarcinoma (ICC) was also performed. MATERIALS AND METHODS: Thirty classical HCCs, 30 ICCs and 10 AFP-producing gastric carcinomas were analyzed. RESULTS: Only one case of HCC had focal labeling of SMARCB1 in the nuclei. Twelve cases of ICC were immunopositive for SMARCB1, with either focal or diffuse reactivity. All AFP-producing gastric carcinomas were immunopositive for SMARCB1 in the nuclei, and the reactivity was consistently diffuse. CONCLUSION: SMARCB1 is a potential marker for distinguishing metastatic AFP-producing gastric carcinoma from HCC.

Diagnostic Utility of Prostein, Uroplakin II and SATB2 for Diagnosing Carcinoma of Unknown Primary Origin: A Systematic Immunohistochemical Profiling.

BACKGROUND/AIM: Immunohistochemistry was used to evaluate 600 carcinomas of major histological types from various organs to determine the tissue distributions of the novel markers prostein, uroplakin II and SATB2. MATERIALS AND METHODS: We retrieved 30 cases from 20 different carcinomas of systemic organs. RESULTS: All prostate adenocarcinomas were immunopositive for prostein, and its reactivity was consistently diffuse. There was faint labeling of prostein in few cases of the 570 non-prostatic carcinomas. Uroplakin II was immunopositive in 53% and 60% of urothelial carcinomas (UC) of the bladder and the ureter, respectively. There was focal and weak positivity of uroplakin II in a few cases of non-urinary tract carcinomas. SATB2 was frequently positive in adenocarcinomas of the digestive organs, and was also expressed in a minority of the non-colorectal adenocarcinomas. CONCLUSION: Prostein and uroplakin II are immunohistochemical biomarkers of prostate adenocarcinomas and UCs of the urinary tract.

High expression of CD10 in anaplastic thyroid carcinomas.

AIMS: CD10 is an endopeptidase that degrades various bioactive peptides in the extracellular matrix. In addition to enzymatic degradation, it affects multiple intracellular signal transduction pathways. CD10 expression has been extensively studied in human epithelial cancers of numerous organs and sites. However, its presence in thyroid carcinomas, especially in anaplastic thyroid carcinoma (ATC), has not been fully determined. An actual CD10 expression in thyroid lesions including a large series of ATC was evaluated. METHODS AND RESULTS: We examined CD10 by immunohistochemistry (IHC) in 152 thyroid lesions: nine adenomatous goitres (AGs) and 143 tumours, including 47 anaplastic carcinomas. IHC showed diffuse and strong positivity for CD10 in the epithelial components of almost all ATCs. However, epithelia with squamous metaplasia and oncocytic change from AGs, follicular adenomas and differentiated carcinomas had focal CD10 reactivity. Some papillary thyroid carcinomas (PTCs), along with the PTC components of some ATCs, showed CD10 positivity in fibroblast-like stromal cells and fibrous material. CONCLUSION: Our results imply that the CD10 expression pattern depended on the histotypes of thyroid lesions. When possible metastatic tumours and non-epithelial tumours are excluded, high CD10 expression may be useful in determining whether a primary thyroid carcinoma includes an anaplastic component.

Single-stranded DNA (ssDNA) Labeling Index Is Related to Risk Grade in Patients with Gastrointestinal Stromal Tumors.

BACKGROUND/AIM: The main objective of the present study was to evaluate the significance of apoptosis in the Fletcher's risk classification for gastrointestinal stromal tumors (GISTs). MATERIALS AND METHODS: Apoptotic cells were identified by immunostaining for single-stranded DNA (ssDNA). We assigned each GIST to one of four risk groups: very low risk, n=32; low risk, n=53; intermediate risk, n=15; high risk, n=6). RESULTS: The mean ssDNA labeling index for each group was 8.0±44.2, 20.1±86.5, 18.7±38.6 and 5.7±5.7, respectively. Fletcher's risk classification for GISTs correlated significantly with the ssDNA labeling index (p=0.002). CONCLUSION: The ssDNA labeling index and Ki-67 labeling index were the most significant factors corresponding to the risk grade of GISTs. These findings suggest that the ssDNA labeling index might be useful for predicting aggressive biological behavior of GISTs.

Esophagogastric Junction Carcinomas - Discriminating Histological Types Through Immunohistochemistry.

BACKGROUND/AIM: Although differentiating squamous cell carcinoma (SCC) from adenocarcinoma (AC) at the esophagogastric junction (EGJ) is important for the choice of treatment, this can occasionally be difficult with small biopsy specimens. Therefore, the purpose of this study was to determine the most useful immunomarker panel for discriminating between SCC and AC of the EGJ. MATERIALS AND METHODS: We analyzed 15 SCCs and 26 ACs of the EGJ obtained surgically using immunohistochemistry. RESULTS: The sensitivities of p40, p63 and cytokeratin 5/6 were 100% with specificities of 88%, 46% and 81%, respectively, for SCC. The sensitivities of CAM5.2, caudal-type homeobox 2 (CDX2), mucin-5AC (MUC-5AC) and MUC-6 were 100%, 81%, 77% and 85% with specificities of 27%, 100%, 87% and 87% for AC. CONCLUSION: We demonstrated that a two-marker panel of p40 and CDX2 is highly sensitive and specific.

Frequent BRAF V600E and Absence of TERT Promoter Mutations Characterize Sporadic Pediatric Papillary Thyroid Carcinomas in Japan.

Pediatric papillary thyroid carcinoma (PTC) has unique features but requires further genetic investigation. Moreover, there has been increasing concern about the risk for pediatric PTC in Japan after the Fukushima accident. This study aims to evaluate the frequencies of BRAF and TERT promoter mutations and to examine their significance in non-radiation-associated pediatric PTCs in Japan. We enrolled 81 pediatric PTC patients aged ≤20 years. The control group included 91 adult PTCs from patients >20 years old. BRAF and TERT mutations were analyzed by allele-specific-PCR and/or Sanger sequencing. Compared with adult PTCs, pediatric PTCs exhibited larger tumor size, more frequent lymph node metastasis, and less classical histology. The prevalence of BRAF V600E in pediatric PTCs was 54% and significantly lower than that in adults of 85%. In the pediatric PTCs, BRAF V600E was positively associated with older age, classical histology, and the lymph node metastasis but independent from other clinicopathological factors. TERT mutations were identified in 13% of adults and in none of the pediatric PTCs. In conclusion, pediatric PTCs are characterized by more advanced clinicopathological features, lower BRAF V600E frequency, and absence of TERT mutation. The BRAF V600E frequency in this study is similar to the reported BRAF V600E frequency in the ultrasonographically screened pediatric PTCs in Fukushima.

Genetic alterations of differentiated thyroid carcinoma in iodine-rich and iodine-deficient countries.

BRAF V600E mutation, RET rearrangements, and RAS mutations are the common genetic alterations in differentiated thyroid carcinomas derived from follicular thyroid cells. However, the relationship between these alterations and iodine intake is still controversial. To clarify the influence of iodine intake on the occurrence of differentiated thyroid carcinomas, we performed molecular analyses for two differentiated carcinomas, papillary thyroid carcinomas (PTCs) and follicular thyroid carcinomas (FTCs), from an iodine-rich country (Japan) and an iodine-deficient country (Vietnam). We examined 120 PTCs (67 Japanese and 53 Vietnamese) and 74 FTCs (51 Japanese and 23 Vietnamese). We carried out allele-specific polymerase chain reaction (AS-PCR) for BRAF V600E, PCR and direct sequencing for RAS mutations (codon 12, 13, and 61 in NRAS, HRAS, and KRAS), and RT-PCR for RET/PTC1 and RET/PTC3. BRAF V600E was present in 55/67 (82.1%) Japanese PTCs and 44/53 (83%) Vietnamese PTCs. RET/PTC1 was identified in only one PTC from each country, and no samples had RET/PTC3. NRAS mutation was found in 17/51 (33.3%) Japanese FTCs and 4/23 (17.4%) Vietnamese FTCs. NRAS mutation was cited in codon 61 (20 cases) and codon 12 (one case). None of FTCs had KRAS or HRAS mutations. There were no significant differences in the prevalence of BRAF V600E, RET/PTC, or RAS mutations between the two countries. Our study showed no differences in genetic alterations of thyroid cancers from iodine-rich and iodine-deficient countries, possibly suggesting that iodine intake might not affect the genetic alterations of differentiated thyroid cancer.

Immunohistochemical detection of NRAS(Q61R) protein in follicular-patterned thyroid tumors.

The NRAS(A182G) mutation, which results in the NRAS(Q61R) protein, is a major driver mutation in follicular-patterned thyroid neoplasms. Although new immunohistochemistry (IHC) for NRAS(Q61R) is now available, its sensitivity, specificity, and diagnostic utility for thyroid tumors are not yet established. We performed IHC for NRAS(Q61R) and direct sequencing for NRAS codon 61 in 4 thyroid cancer-derived cell lines and 98 follicular-patterned thyroid tumors that included 22 follicular thyroid adenomas (FTAs), 35 follicular thyroid carcinomas (FTCs), and 41 cases of nodular hyperplasia (NH). In the tumors with NRAS(Q61R), the expression of BRAF(V600E) was further evaluated immunohistochemically. Two cell lines with NRAS(A182G) showed selective immunoreactivity for NRAS(Q61R). In tumor tissues, NRAS(Q61R) IHC was positive in 18% (4/22), 29% (10/35), and 2% (1/41) of FTAs, FTCs, and NH samples, respectively. The frequencies of the NRAS(Q61R) in FTAs and FTCs were significantly higher than that in NH (P=.046 and P=.001, respectively). All tumors with NRAS(Q61R) expression exhibited uniform cytoplasmic positivity with or without accumulation in their cell membranes. Of the 15 tumors with NRAS(Q61R) expression, 13 cases showed NRAS(A182G) in direct sequencing, whereas all of the tumors without NRAS(Q61R) expression were negative for the mutation. There were no tumors with overlapping expression of NRAS(Q61R) and BRAF(V600E). In reference to the direct sequencing, sensitivity and specificity of the NRAS(Q61R) IHC were 100% and 98%, respectively. In conclusion, NRAS(Q61R) IHC is a highly sensitive and specific tool that is useful for differentiating follicular-patterned thyroid tumors.