Peter Shintaku

Primary Glioblastomas Express Mesenchymal Stem-Like Properties

Glioblastoma is the most common and aggressive primary brain cancer. Recent isolation and characterization of brain tumor-initiating cells supports the concept that transformed neural stem cells may seed glioblastoma. We previously identified a wide array of mesenchymal tissue transcripts overexpressed in a broad set of primary glioblastoma (de novo) tumors but not in secondary glioblastoma (derived from lower-grade) tumors, low-grade astrocytomas, or normal brain tissues. Here, we extend this observation and show that a subset of primary glioblastoma tumors and their derived tumor lines express cellular and molecular markers that are associated with mesenchymal stem cells (MSC) and that glioblastoma cell cultures can be induced to differentiate into multiple mesenchymal lineage-like cell types. These findings suggest either that a subset of primary glioblastomas derive from transformed stem cells containing MSC-like properties and retain partial phenotypic aspects of a MSC nature in tumors or that glioblastomas activate a series of genes that result in mesenchymal properties of the cancer cells to effect sustained tumor growth and malignant progression. (Mol Cancer Res 2006;4(9):607–19)

Her-2/neu gene amplification in low to moderately expressing breast cancers: possible role of chromosome 17/Her-2/neu polysomy.

Overexpression of the Her‐2/neu (HER2) oncogene is known to confer important prognostic and predictive value to patients with breast cancer. Controversy exists as to the best method for its determination caused primarily by the variable sensitivities of the different antibodies and interobserver differences, particularly in the group of breast cancers with borderline levels of expression of the protein product. This study was therefore designed to determine the status of the HER2 gene amplification in a group of breast carcinomas with low levels of overexpression. After an initial validation of our procedures, a series of 52 consecutive cases of formalin‐fixed, paraffin‐embedded breast cancers with low levels of overexpression and a series of 22 cases with no expression by immunohistochemistry were analyzed by fluorescence in situ hybridization (FISH), and the results correlated statistically. Amplification of the HER2 gene was observed in 16% of equivocal to weakly positive cases. Those that were amplified showed low levels of amplification with ratios less than 4.5 and a characteristic scattered pattern of distribution of HER2 signals in the FISH assay. In addition, heterogeneity was noted in two cases in the amplification of the HER2 gene within the same tumor samples with pockets of amplified tumor cells amidst nonamplified tumor cells. In cases without amplification, a statistically significant number showed chromosome 17 polysomy. In conclusion, equivocal to low levels of HER2 overexpression in breast cancers are associated, in the majority of cases, with chromosome 17 polysomy and a corresponding increase in the HER2 gene numbers. True gene amplification is present in only a minority of cases. FISH analysis should be used for confirmation of gene amplification. Prior screening and selection of appropriate immunohistochemistry‐positive areas for FISH analysis may prove beneficial.

Immunoperoxidase staining for C4d on paraffin-embedded tissue in cardiac allograft endomyocardial biopsies: comparison to frozen tissue immunofluorescence.

C4d deposition in microvasculature is a marker for humoral rejection. The authors compared a recently developed C4d immunoperoxidase (IP) method for paraffin-embedded tissue to immunofluorescence (IF) of frozen tissue. Of 315 frozen endomyocardial biopsies with IF staining for C4d, 280 were negative and 35 were positive. Negative controls were 17 negative biopsies and 11 biopsies with myocyte necrosis. The extent of IP and IF staining was graded as 0 to 3+. Staining intensity and the number and type of positive vessels were recorded. Staining patterns in Quilty lesions (QL) and foci of acute cellular rejection (ACR) were also evaluated. In 34 biopsies with sufficient tissue, IP criteria of 2+/3+, or more than 10 to 20 positive vessels per 10 high-power fields detected 25.0% (1/4), 18.2% (2/11), and 84.2% (16/19) of 1+, 2+, and 3+ IF-positive biopsies, respectively, without false positives. Considering C4d IF 3+ as positive resulted in 84.2% (16/19) sensitivity and 93.0% specificity (40/43). Intensely stained capillaries predominated in six of seven biopsies when more than 100 capillaries per 10 high-power fields were positive. Seventy percent (7/10) of IP 2+ and 3+ biopsies showed positive capillaries in QLs, while 36.4% (4/11) of IP 1+ and negative biopsies did. All eight IP 2+/3+ biopsies showed positive capillaries in ACR foci, while 25.0% (1/4) of IP-negative biopsies did. Capillary staining in QLs and areas of ACR reflects overall C4d deposition. In conclusion, IP staining of 2+/3+ is highly sensitive and specific for C4d positivity. The authors recommend considering 2+ and 3+ as positive staining when using the IP technique.

Mucinous adenocarcinomas of the thymus : Report of 2 cases and review of the literature

BackgroundMost adenocarcinomas of the mediastinum are metastatic lesions. Primary thymic adenocarcinomas are extremely rare neoplasms. We could find only 12 cases reported in the literature; of these 12, only 4 were of the mucinous subtype. DesignWe report 2 additional cases of the mucinous subtype, including a previously unreported mucinous variant with numerous psammoma bodies. ResultsThe first case in a 61-year-old woman resembled a mucinous (colloid) carcinoma of other organs such as the breast and colon. It consisted of islands and strips of tumor cells floating in large pools of extracellular mucin. A unique feature of this tumor was the presence of numerous psammoma bodies. Immunohistochemically, the tumor cells were positive for cytokeratin (CK) 7 and negative for CD5. The second case in an 82-year-old woman was a mucinous adenocarcinoma arising from a thymic cyst with areas of transition from benign to dysplastic epithelium. The tumor cells formed dilated glands, cords, and small nests that infiltrated the thymic cyst wall and exhibited evidence of mucin production. Immunohistochemically, the tumor cells were positive for CK 7 and focally positive for both CD5 and CK 5/6. ConclusionsMucinous adenocarcinoma, with or without, psammoma bodies, may be of primary thymic origin and should be considered in the differential diagnosis of malignant mediastinal tumors. These 2 cases provide further documentation of the rare occurrence of primary mucinous adenocarcinomas of the thymic gland.

Immunohistochemical Analysis of Lung Carcinomas With Pure or Partial Bronchioloalveolar Differentiation

CONTEXT In 1999, the World Health Organization redefined bronchioloalveolar carcinomas (BACs) as those neoplasms with only a pure lepidic growth pattern and no invasion. OBJECTIVES The present study examined 45 lung cancers with a BAC component (1) to determine whether these tumors would be classified as BACs by current World Health Organization standards, (2) to quantitate the BAC component within these tumors, and (3) to see if phenotypic differences exist between the so-called invasive and noninvasive regions of these tumors. DESIGN Retrospective review of hematoxylin-eosin-stained slides and classification of histologic grade, tumor subtype, and percentage of pure BAC pattern, with further characterization by immunohistochemical staining for thyroid transcription factor 1, cytokeratin 7, cytokeratin 20, and Ki-67 antibodies. RESULTS Only 7 (15.6%) of the 45 tumors examined could be classified as BAC by current strict World Health Organization criteria. Those tumors, classified as nonmucinous and mixed, showed similar immunohistochemical staining for cytokeratin 7, cytokeratin 20, and thyroid transcription factor 1; mucinous tumors showed disparate staining. Significant differences in immunohistochemical staining and tumor cell proliferation were seen for the regions of tumors designated as lepidic, infiltrative, and leading edge and for the regions of tumors with different histologic grades (ie, well, moderately, and poorly differentiated). CONCLUSIONS Nonmucinous and mixed BACs are phenotypically similar and show identical immunohistochemical staining patterns; mucinous tumors, on the other hand, show disparate immunohistochemical staining. Pulmonary neoplasms designated as adenocarcinomas with a BAC component represent a heterogenous group with a range of cell types, differentiation, growth, and immunophenotypes. Within an individual neoplasm, there are regional differences in these parameters as well.

Involucrin in intraepithelial and invasive squamous cell carcinomas of the cervix: An immunohistochemical study

Immunohistochemical staining for involucrin, a cytoplasmic protein synthesized during squamous maturation, was assessed in histologic sections from hysterectomy and cone biopsy specimens from patients with cervical neoplasia. In normal and condylomatous squamous epithelium, diffuse cytoplasmic staining was seen in the suprabasal layers, with no staining of the basal cells. Staining was absent in two cases of cervical intraepithelial neoplasia (CIN), grade III, in which the lesions were composed entirely of undifferentiated cells and markedly decreased in cases involving large numbers of basal cells. In 19 of 23 cases (83 per cent) of CIN, however, focal staining for involucrin was seen in large differentiated cells in the more superficial layers, and in two cases of keratinized CIN diffuse suprabasal staining was observed. Similarly, strong staining for involucrin was present in differentiated areas in one case of microinvasive squamous cell carcinoma and in 93 per cent of cases of infiltrating squamous cell carcinoma. These findings suggest that involucrin is a marker for maturation in cervical squamous epithelial neoplasms. Patterns of immunohistochemical staining for involucrin in keratinized dysplasia and differentiated squamous carcinomas should be taken into consideration if loss of involucrin staining is used as a criterion for neoplastic transformation of cervical epithelium, as has been proposed.

Comparison of benign keratoses using p53, bcl‐1, and bcl‐2

While cell‐cycle markers have been used to differentiate benign vs. malignant lesions and to classify malignant lesions, benign keratoses have not been well studied using such markers. We hypothesized that inflammation or irritation of benign keratoses may be related to a shift in the cell cycle. We compared the immunohistochemical staining patterns of 10 seborrheic keratoses (SKs), 10 inflamed seborrheic keratoses (iSKs), and 10 inverted follicular keratoses (IFKs) using antibodies to p53, bcl‐1, and bcl‐2. Staining with antibodies to p53 was slightly increased in IFKs compared with iSKs or non‐inflamed seborrheic keratoses. Bcl‐1 staining was similar in all lesions. A population of bcl‐2‐positive dendritic cells was seen within the epidermal portion of IFKs. Keratinocyte bcl‐2 staining was significantly higher in SKs compared with the other two keratoses. Bcl‐2 may be increased in SKs as an anti‐apoptotic mechanism.

Programmed death-ligand 1 (PD-L1) is expressed in a significant number of the uterine cervical carcinomas

BackgroundThe programmed death-1/programmed death-ligand-1 (PD-1/PD-L1) immune regulatory axis has emerged as a promising new target for cancer therapeutics, with lasting responses seen in the treatment of metastatic renal and lung carcinomas, as well as melanomas. As tumor surface expression of PD-L1 has been found to correlate with objective responses to anti-PD-L1 immunotherapies, we investigated the expression of PD-L1 in human cervical tumors and provide an adopted scoring system for the systematic evaluation of PD-L1 staining.MethodsImmunohistochemical staining for PD-L1 expression was performed on a tissue microarray of 101 normal and neoplastic cervical tissues. Neoplastic cores were divided into three groups: squamous cell carcinoma, adenosquamous carcinoma, and endocervical adenocarcinoma. PD-L1 expression was scored based on an adopted scoring system accounting to percentage and intensity of positivity, and results provided alongside available clinical and demographic data.ResultsOverall, PD-L1 was positive in 32 of 93 (34.4%) cervical carcinomas. Subcategorically, PD-L1 was positive in 28 of 74 (37.8%) squamous cell carcinomas, two of seven (28.6%) adenosquamous carcinomas, and two of 12 (16.7%) endocervical adenocarcinomas. It was negative in six benign cervical tissues.ConclusionsThis study shows a significant expression of PD-L1 in 34.4% of cervical carcinomas and no expression of PD-L1 in benign cervical tissues. These findings suggest a role for further investigation of anti-PD-L1/PD-1 immunotherapies in the treatment of PD-L1-positive cervical tumors. In addition, our adopted scoring system will facilitate more systematic correlations between tumor reactivity and response to treatment.

An infant with MLH3 variants, FOXG1-duplication and multiple, benign cranial and spinal tumors: A clinical exome sequencing study.

A 4‐month‐old male infant presented with severe developmental delay, cerebellar, brainstem, and cutaneous hemangiomas, bilateral tumors (vestibular, hypoglossal, cervical, and lumbar spinal), and few café‐au‐lait macules. Cerebellar and lumbar tumor biopsies revealed venous telangiectasia and intraneural perineuroma, respectively. Sequencing NF1, NF2, and RASA1 (blood), and NF2 and SMARCB1 (lumbar biopsy) was negative for pathogenic mutations. Clinical exome sequencing (CES), requested for tumor syndrome diagnosis, revealed two heterozygous missense variants, c.359T>C;p.Phe120Ser and c.3344G>A;p.Arg1115Gln, in MLH3 (NM_001040108.1), a DNA mismatch repair (MMR) gene, Polyphen‐predicted as probably damaging, and benign, respectively. Sanger sequencing confirmed both variants in the proband, and their absence in the mother; biological father unavailable. Both biopsied tissues were negative for microsatellite instability, and expressed MLH1, MSH2, PMS2, MSH6, and MLH3 immunohistochemically. Chromosomal microarray showed a 133 kb segment copy number duplication of 14q12 region encompassing FOXG1, possibly explaining the developmental delay, but not the tumors. The presence of MLH3 variants with multiple benign neural and vascular tumors was intriguing for their possible role in the pathogenesis of these neoplasms, which were suspicious for, but not diagnostic of, constitutional MMR deficiency. However, functional assays of non‐neoplastic patient‐derived cells showed intact base‐base MMR function. Also, no previous FOXG1‐aberrant patient was reported with tumors. We now report a 3‐year‐old FOXG1‐duplicated patient with a yet undescribed tumor syndrome with clinical features of neurofibromatosis types I and II, where several validation studies could not ascertain the significance of CES findings; further studies may elucidate precise mechanisms and diagnosis for clinical management, including tumor surveillance.

Evaluation of PAX8 Expression in Brain Tissue and Related Neoplasms.

Undifferentiated brain tumors represent a diagnostic challenge, particularly in small biopsies, with regards to their primary versus metastatic origin. The latter may show overlapping morphologic features with primary high-grade brain tumors. In recent years several new antibodies have entered the realm of daily pathology practice. PAX8 (mammalian paired box genes 1 to 9 protein encoding gene) is among these new markers and is recognized as a differentiating marker of the primary site in epithelial tumors outside of the central nervous system. A review of the literature shows lack of site-specific studies with regards to the expression of PAX8 in the central nervous system and its neoplasms. Using this marker we investigated its immunohistochemical expression in normal brain tissue and glial tumors. The immunostain was performed on tissue microarrays of 71 cores from 24 cases. We also performed PAX8 immunostain on sections from cerebellum, pons, periventricular ependymal layer, choroid plexus, pituitary, and meninges of 3 autopsy cases. Our results indicate lack of PAX8 expression by benign brain tissue. Only 1 glioblastoma core (1/9 cores) showed focal nuclear reactivity with the antibody. Our results indicate that presence of PAX8 immunoreactivity in an undifferentiated brain tumor lacking gliofibrillary acidic protein expression should prompt consideration of a metastatic tumor.