Histopathologic reassessment of melanoma and other melanocytic skin lesions excised in 2009 and 2018-2019. / Histopatologisk revurdering av melanom og andre melanocytære hudlesjoner eksidert i 2009 og 2018­19.

BACKGROUND: Histopathological assessment of melanoma and other melanocytic skin lesions can be difficult and can vary between pathologists. MATERIAL AND METHOD: Histopathological slides of 196 melanocytic skin lesions from 2009 and 2018-2019 were obtained from the archive of the Department of Pathology at Oslo University Hospital and classified into six diagnostic categories: 1) benign nevus, 2) irregular/dysplastic nevus, i.e. dysplastic nevus with moderate atypia, 3) nevus with severe atypia, i.e. dysplastic nevus with severe atypia, 4) melanoma in situ, 5) superficial spreading or lentiginous melanoma and 6) nodular melanoma. The slides were then examined independently and blindly by three experienced pathologists and categorised in the same way. Interobserver agreement was assessed with Cohen's kappa, and agreement with the original diagnosis was assessed by the proportion of assessments in the same diagnostic category. RESULTS: The kappa values for the assessments from the three pathologists ranged from 0.45 to 0.50. The proportion of reassessments in agreement with the original diagnostic category was 85.7 % (95 % CI 75.7 to 92.1), 29.2 % (19.9 to 40.5), 27.8 % (20.9 to 36.0), 78.3 % (70.4 to 84.5), 81.2 % (73.7 to 86.9) and 93.3 % (82.1 to 97.7), respectively, i.e. highest for nodular melanoma. The proportion of reassessments in which the diagnosis was more serious or less serious than the original diagnosis was higher and lower, respectively, for slides from 2009 than for slides from 2018-2019. INTERPRETATION: The differences between the pathologists' assessments and deviations from the original diagnoses can be explained by poorly reproducible diagnostic criteria, diagnostic entities with overlapping morphology and increasing awareness of early signs of malignancy. Some evolution in diagnostic practice cannot be ruled out.

MX 2 is a novel regulator of cell cycle in melanoma cells.

MX2 protein is a dynamin-like GTPase2 that has recently been identified as an interferon-induced restriction factor of HIV-1 and other primate lentiviruses. A single nucleotide polymorphism (SNP), rs45430, in an intron of the MX2 gene, was previously reported as a novel melanoma susceptibility locus in genome-wide association studies. Functionally, however, it is still unclear whether and how MX2 contributes to melanoma susceptibility and tumorigenesis. Here, we show that MX2 is differentially expressed in melanoma tumors and cell lines, with most metastatic cell lines showing lower MX2 expression than primary melanoma cell lines and melanocytes. Furthermore, high expression of MX2 RNA in primary melanoma tumors is associated with better patient survival. Overexpression of MX2 reduces in vivo proliferation partially through inhibition of AKT activation, suggesting that it can act as a tumor suppressor in melanoma. However, we have also identified a subset of melanoma cell lines with high endogenous MX2 expression where downregulation of MX2 leads to reduced proliferation. In these cells, MX2 downregulation interfered with DNA replication and cell cycle processes. Collectively, our data for the first time show that MX2 is functionally involved in the regulation of melanoma proliferation but that its function is context-dependent.

Fusion of the Genes PHF1 and TFE3 in Malignant Chondroid Syringoma.

BACKGROUND/AIM: Malignant chondroid syringoma is a rare tumor of unknown pathogenesis. MATERIALS AND METHODS: Genetic analyses were performed on a malignant chondroid syringoma. RESULTS: G-banding analysis of short-term cultured tumor cells yielded the karyotype 46,Y,t(X;6)(p11;p21)[15]/46,XY[2]. RNA sequencing detected an in-frame fusion of PHF1 from 6p21 with TFE3 from Xp11, verified by RT-PCR and Sanger sequencing. Genomic PCR showed that the PHF1-TFE3 junction was identical to the fusion found by RNA sequencing and RT-PCR. CONCLUSION: Malignant chondroid syringoma is genetically related to tumors with PHF1 rearrangements such as low-grade endometrial sarcoma and ossifying fibromyxoid tumor, but also with tumors having TFE3 rearrangements such as renal cell carcinoma, alveolar soft part sarcoma, PEComa, and epithelioid hemangioendothelioma. Further investigations on malignant chondroid syringomas are needed in order to determine whether genetic heterogeneity exists among them and the clinical impact of the PHF1-TFE3 fusion.

Melanoma staging: Varying precision and terminal digit clustering in Breslow thickness data is evident in a population-based study.

BACKGROUND: Errors in Breslow thickness reporting can give misclassification of T category, an important classifier in melanoma staging. OBJECTIVE: We sought to investigate precision (number of digits) and terminal digit clustering in Breslow thickness and potential consequences for T category. METHODS: All first primary and morphologically verified invasive melanomas in Norway between 2008 and 2015 were included. A smoothing model was fitted to estimate the underlying Breslow thickness distribution without digit clustering. RESULTS: Thickness was reported for 13,057 (97.5%) patients; the median was 1.0 mm (range, 0.09-85). It was reported as whole numbers (15.6%), to 1 decimal (78.2%) and 2 decimal places (6.2%)-thin tumors with more precision than thick tumors. Terminal digit clustering was found with marked peaks in the observed frequency distribution for terminal digits 0 and 5, and with drops around these peaks. Terminal digit clustering increased proportions of patients classified with T1 and T4 tumors and decreased proportions classified with T2 and T3. LIMITATIONS: Breslow thickness was not reported in 2.5% of cases. CONCLUSIONS: The Norwegian recommendation of measurement to the nearest 0.1 mm was not followed. Terminal digit clustering was marked, with consequences for T category. Pathologists, clinicians, and epidemiologists should know that clustering of thickness data around T category cut points can impact melanoma staging with consequent effect on patient management and prognosis.

Fusion of the genes ataxin 2 like, ATXN2L, and Janus kinase 2, JAK2, in cutaneous CD4 positive T-cell lymphoma.

Acquired mutations were recently described in cutaneous T-cell lymphomas for the JAK1, JAK3, STAT3, and STAT5B genes of the JAK-STAT pathway. In the present study, RNA-sequencing of a primary cutaneous CD4 positive T-cell lymphoma carrying a three-way t(9;13;16)(p24;q34;p11) chromosome translocation showed that JAK2 from chromosome band 9p24 was rearranged and fused to a novel partner gene, ATXN2L, from 16p11. RT-PCR together with Sanger sequencing verified the presence of the ATXN2L-JAK2 fusion transcript. The ATXN2L-JAK2 fusion gene would code for a chimeric protein containing all domains of ATXN2L and the catalytic domain of the JAK2 tyrosine kinase. The ATXN2L-JAK2 chimeric protein could lead to constitutive activation of the downstream JAK-STAT signaling pathway in a manner similar to that seen for other JAK2 fusion proteins.

Combination of Juvenile Papillomatosis, Juvenile Fibroadenoma and Intraductal Carcinoma of the Breast in a 15-Year-old Girl.

UNLABELLED: The association of juvenile papillomatosis with carcinoma is proven, but very rare, as there exist several reported cases. CASE REPORT: A 15-year-old girl with no family history of breast cancer presented with two masses in the left breast. The excisional biopsy on both lumps revealed juvenile fibroadenoma and juvenile papillomatosis epithelial proliferation multiple cystic expanded channels. In some of these channels, cytological features of intraductal carcinoma were observed. We performed a full immunohistochemical examination of the juvenile papillomatosis. The patient refused any further surgical or adjuvant treatment. There are no signs of recurrence in the 15 year follow-up. DISCUSSION: This case is a diagnostic and therapeutic challenge, taking into account the patient's age and the controversial treatment recommendations. Good collaboration between surgeons and pathologists is essential for an accurate diagnostic process and aims to avoid under- or overtreatment.

Lymphoplasmacytic lymphoma and marginal zone lymphoma in the bone marrow: paratrabecular involvement as an important distinguishing feature.

OBJECTIVES: The differential diagnosis between bone marrow involvement by lymphoplasmacytic lymphoma (LPL) and marginal zone lymphoma (MZL) is challenging because histology and immunophenotype of both diseases overlap. We revisited the diagnostic pathology features of both diseases in the bone marrow. METHODS: We studied a series of bone marrow trephine biopsy specimens from 59 patients with Waldenström macroglobulinemia without extramedullary involvement and bone marrow biopsy specimens from 23 patients with well-characterized MZL who also had bone marrow involvement. H&E- and immunoperoxidase-stained sections of bone marrow trephine biopsy specimens as well as flow cytometry and classic cytogenetics performed on aspirations were reviewed. The study was complemented with MYD88 L265P mutation analysis of all samples. RESULTS: The most distinguishing features of LPL with respect to MZL were focal paratrabecular involvement (P < .001), the presence of lymphoplasmacytoid cells (P < .001) and Dutcher bodies (P < .001), increased numbers of mast cells (P < .001), and the MYD88 L265P mutation (P < .001). CONCLUSIONS: LPL can be reliably distinguished from MZL in the bone marrow by using a combination of pathology characteristics. Our findings stress the diagnostic importance of using the combination of the following parameters for a correct LPL diagnosis: paratrabecular infiltration, the presence of lymphoplasmacytoid cells and cells with Dutcher bodies, and an increased number of mast cells in addition to the presence of MYD88 mutation.

Clinical and histopathological features of folliculotropic mycosis fungoides: a Norwegian patient series.

Folliculotropic mycosis fungoides is a variant of cutaneous T-cell lymphoma with distinct clinicopathological features. We describe here the clinical presentation, pathology findings and treatment outcome in 15 Norwegian patients. All patients were diagnosed between 1997 and 2010 at Oslo University Hospital. A spectrum of skin lesions, both typical and atypical, such as leonine facies, acneiform lesions, psoriasiform plaques, purulent ulcerations and cystic milia-like lesions for mycosis fungoides, were seen. Histological examination revealed characteristic infiltration of hair follicles with neoplastic T cells associated with partial destruction of the former. A CD4+ immunophenotype of the neoplastic T cells with loss of one or more T-cell markers was demonstrated. In general, the patients were given more aggressive therapeutic regimens than those with conventional mycosis fungoides, and showed a trend towards more rapid disease progression. In conclusion, this case series confirms the distinct clinical and histological features of folliculotropic mycosis fungoides.

Expression of cyclooxygenase-2 in invasive breast carcinomas and its prognostic impact.

Representative tumour sections from 468 patients with invasive breast cancer were immunostained for cyclooxygenase-2 (COX-2) and evaluated. The relationships between COX-2 expression, clinical outcome and various clinicopathological variables, including tumour vascularity and disseminated tumour cells (DTC) in the bone marrow were examined. COX-2 expression in invasive breast carcinoma cells was positively associated with oestrogen receptor and/or progesterone receptor positivity (p<0.001). Triple-negative tumours showed no/low COX-2 expression more frequently than other tumour types (p<0.001). Expression of COX-2 was not associated with breast cancer-specific survival (p=0.49, log-rank) or distant disease-free survival (p=0.67, log-rank) for all patients, including lymph node-negative, untreated patients (p>0.14, log-rank). There was also no significant association between COX-2 expression and histological grade, tumour size, nodal status, DTC in bone marrow, p53, HER2, or tumour vascularity. In conclusion, COX-2 expression in this series was associated with the presence of hormone receptors. Low COX-2 expression was observed in triple-negative breast carcinomas.

Expression of vascular endothelial growth factor and vascular endothelial growth factor receptors 1 and 2 in invasive breast carcinoma: prognostic significance and relationship with markers for aggressiveness.

AIMS: Vascular endothelial growth factor (VEGF), VEGF receptor 1 (VEGFR-1) and VEGF receptor 2 (VEGFR-2) play a role in breast cancer growth and angiogenesis. We examined the expression and relationship with clinical outcome and other prognostic factors. METHODS AND RESULTS: Tumour sections from 468 breast cancer patients were immunostained for VEGF, VEGFR-1, and VEGFR-2, and their relationships with tumour vascularity, disseminated tumour cells (DTCs) in bone marrow and other clinicopathological parameters were evaluated. VEGF, VEGFR-1 and VEGFR-2 immunoreactivities were observed in invasive breast carcinoma cells. VEGF expression was significantly associated with VEGFR-1 and VEGFR-2 expression (P < 0.001). High-level cytoplasmic expression of VEGFR-1 was associated with significantly reduced distant disease-free survival (DDFS) (P = 0.017, log-rank) and breast cancer-specific survival (BCSS) (P = 0.005, log-rank) for all patients, and for node-negative patients without systemic treatment (DDFS, P = 0.03, log-rank; BCSS, P = 0.009, log-rank). VEGFR-1 expression was significantly associated with histopathological markers of aggressiveness (P < 0.05). Significantly reduced survival was observed in DTC-positive patients as compared with DTC-negative patients in the combined moderate/high VEGFR-1 group (P < 0.001 for DDFS and BCSS), and the same was true for DDFS in the moderate VEGFR-2 group (P = 0.006). CONCLUSIONS: High-level expression of VEGFR-1 indicates reduced survival. Higher-level expression of VEGFR-1 or VEGFR-2 in primary breast carcinomas combined with the presence of DTC selects a prognostically unfavourable patient group.

Transformation of B cell lymphoma to histiocytic sarcoma: somatic mutations of PAX-5 gene with loss of expression cannot explain transdifferentiation.

Transdifferentiation of B cell lymphoma of germinal center cell origin to histiocytic sarcoma has recently been described but is a rare occurrence. The cause for loss of B cell differentiation in these lymphomas is unknown. We investigated whether somatic hypermutation of the PAX-5 gene, a transcription factor that is important for maintaining B cell identity and is frequently mutated in B cell lymphomas of germinal center cell origin, might be a cause for loss of PAX-5 expression and thus B cell phenotype. However, no somatic hypermutation of the PAX-5 gene was detected in the two cases we studied. The molecular basis for transdifferentiation of B cell lymphoma to histiocytic sarcoma remains therefore unresolved.

Breast carcinoma vascularity: a comparison of manual microvessel count and Chalkley count.

Manual counting of microvessels as intratumoral microvessel density (MVD) and Chalkley counting have been used in several studies to assess the prognostic impact of vascularity in invasive breast carcinomas. In our present study, the aim was to evaluate the prognostic value of angiogenesis in invasive breast carcinoma assessed by MVD and Chalkley techniques in the same series of patients. A total of 498 breast carcinoma patients with median follow up time 85 months were evaluated. The tumour vascularity was quantified by both manual microvessel count (MVD) and Chalkley count in CD34 stained breast carcinoma slides by a single investigator blinded to clinical information. Other relevant clinicopathological parameters were noted, including breast cancer related death and both loco-regional and systemic relapse. The patients were stratified by converting MVD and Chalkley counts to categorical variables to assess prognostic impact, and results were compared. High vascular grades using MVD count did not demonstrate any prognostic significance for breast cancer specific survival (BCSS) or distant disease free survival (DDFS) either in whole patient group (BCSS, p=0.517, DDFS, p=0.301) or in non-treated node negative patients (p>0.05). Chalkley count showed prognostic significance for both DDFS and BCSS in whole patient group (p<0.001) and also in untreated node negative patient group (p<0.05). In multivariate analysis, Chalkley count, but not MVD, retained the prognostic value for BCSS (p=0.007) and DDFS (p=0.014). The Chalkley count for assessing angiogenesis in invasive breast carcinomas demonstrated prognostic value. The Chalkley method appears to be the better method in estimating the prognostic impact of vascularity in invasive breast carcinomas.

Mantle cell lymphoma with partial involvement of the mantle zone: an early infiltration pattern of mantle cell lymphoma?

Most patients with mantle cell lymphoma present with a diffuse or nodular infiltration of the involved organs at diagnosis. We present two patients with a rare morphological variant, displaying a partial involvement of the mantle zone. Patient 1 presented with an enlarged inguinal lymph node, which showed marked follicular hyperplasia with singly spread Cyclin D1+ small lymphoid cells in the mantle zones. An additional lymph node biopsy taken 3 months later showed the same pattern. Patient 2 presented with a classical mantle cell lymphoma with lymph node, bone marrow and gastro-intestinal involvement. However, revision of an appendectomy specimen taken 4 years earlier showed pronounced follicular hyperplasia with singly spread Cyclin D1+ small lymphoid cells in the mantle zones. Mantle cell lymphoma with partial involvement of the mantle zone has rarely been reported and many represent an early manifestation of mantle cell lymphoma. Our cases also illustrate that the inclusion of an anti-cyclin D1 antibody in the diagnostic panel of antibodies to study unexplained follicular hyperplasia, might be advised.

Vascularization in primary breast carcinomas: its prognostic significance and relationship with tumor cell dissemination.

PURPOSE: The interaction between tumor cells, stroma, and endothelial cells is important for the dissemination of tumor cells. The aim of the present study is to examine vascularity in primary breast carcinomas and its prognostic significance and relationship with tumor cell dissemination. EXPERIMENTAL DESIGN: A total of 498 invasive breast carcinomas were analyzed. Representative tumor sections were stained for CD34 and CD105, and vascularity was quantified by the Chalkley method. The relationship between Chalkley counts, vascular invasion, disseminated tumor cells (DTC) in the bone marrow, other clinicopathologic variables, and clinical outcome was evaluated. RESULTS: High vascular grades determined by Chalkley counts were significantly associated with shorter distant disease-free survival and breast cancer-specific survival in all patients (P < 0.001, log-rank) and in node-negative patients not receiving adjuvant systemic therapy (P < 0.05). In multivariate analysis, both CD34 and CD105 Chalkley counts showed prognostic significance for distant disease-free survival (P = 0.014 and P = 0.026), whereas CD34 also showed prognostic significance for breast cancer-specific survival (P = 0.007). Vascular invasion and DTCs in the bone marrow showed independent prognostic significance. DTC did not discriminate survival for CD34 low Chalkley counts, whereas a very poor prognosis was observed for DTC-positive patients with high CD34 counts. In node-negative patients not receiving systemic chemotherapy, high CD34 and high CD105 counts in combination identified patients with unfavorable outcome, as opposed to all other CD34/CD105 combinations. CONCLUSIONS: Improved identification of risk groups could be obtained by adding CD34 and CD105 vascular analysis to DTC, vascular invasion, and other primary tumor factors. This may facilitate the selection of candidates for adjuvant systemic therapy.

D2-40 is not a specific marker for cells of mesothelial origin in serous effusions.

The presence of effusion in a patient with a history of primary malignant tumor elsewhere in the body is generally accepted as a clinical manifestation of metastatic disease. Even in those cases, it is sometimes difficult to differentiate reactive mesothelial cells from carcinoma cells. Another challenging issue especially in the field of serous effusions is the differential diagnosis between malignant mesothelioma and metastatic adenocarcinoma. The aim of this study was to evaluate the potential use of the D2-40 antibody detecting the M2A oncofetal antigen in the diagnosis of malignant serous effusions. Two hundred and ninety effusion specimens (169 ovarian carcinomas, 44 breast carcinomas, 32 malignant mesotheliomas, 6 lung carcinomas, 8 reactive specimens, and 31 tumors of other origin) were assessed. Expression in reactive mesothelial cells was additionally assessed on 145 malignant effusions. Immunohistochemical analysis using the EnVision system was performed. M2A antigen was expressed in malignant mesotheliomas and reactive mesothelial cells in all specimens. Positive membranous staining was observed in 58% of ovarian carcinomas, 33% of lung carcinomas, and 30% of breast carcinomas. Pulmonary, breast, and nonovarian gynecologic tumors usually showed weak focal membranous staining, whereas the ovarian adenocarcinomas showed an expression pattern more similar to mesotheliomas. The results from the present study suggest low specificity for D2-40 as a mesothelial marker, especially in the context of differentiating mesothelial cells from ovarian carcinoma, and argue against its inclusion in the diagnostic panel of serous effusions.

The mitogen-activated protein kinases (MAPK) p38 and JNK are markers of tumor progression in breast carcinoma.

OBJECTIVE: To investigate the activation of mitogen-activated protein kinases (MAPK) in breast carcinoma effusions and to analyze its relationship to anatomic site and clinical parameters. METHODS: Activated MAPK (p-ERK, p-JNK, and p-p38) expression was studied in 42 effusions and 51 corresponding solid tumors (23 primary, 28 metastases) using immunohistochemistry (IHC). Hormone receptor and HER2 status, proliferation (Ki-67), and apoptosis (p85-PARP fragment) were assessed. MAPK levels, activity, and activation ratio (phospho/pan-MAPK ratio) were analyzed in 19 effusions using immunoblotting (IB). RESULTS: Nuclear expression of p-p38 and p-JNK was significantly higher in effusions compared with both primary tumors (P < 0.001 for p-JNK, P = 0.011 for p-p38) and lymph node metastases (P = 0.003 for p-JNK, P = 0.025 for p-p38) but was not accompanied by apoptosis. IB showed pan-ERK and p-ERK in 18/19 effusions, pan-JNK and p-JNK in 18/19 and 17/19 effusions, respectively, and pan-p38 and p-p38 in 19/19 and 17/19 specimens, respectively. In univariate survival analysis of all cases, advanced disease stage (P = 0.041), previous chemotherapy (P = 0.004), and radiation (P = 0.001) and higher Ki-67 scores (P = 0.01) correlated with worse overall survival (OS). In Cox multivariate analysis, stage (P = 0.018), chemotherapy (P = 0.024), radiation (P = 0.017), and ER status (P = 0.002) were independent prognosticators of OS. Quantitative analysis of IB data showed that higher p38 activation ratio correlates with shorter OS (P = 0.01). CONCLUSIONS: This study presents first evidence of in vivo activation of MAPK in breast carcinoma effusions. The elevated JNK and p38 activation in effusions may be a stress-related mechanism providing breast carcinoma cells with survival advantages rather than a drive towards apoptosis. p38 and Ki-67 may be new prognostic markers for patients with breast cancer effusions.

Genomic aberrations in mucinous tubular and spindle cell renal cell carcinomas.

Mucinous tubular and spindle cell carcinoma of the kidney is a new diagnostic entity. We present the pathologic and genomic characteristics of three such low-malignant tumors. Two of the tumors were found in women aged 19 and 52 years, the third tumor was found in an 80-year-old man, and the tumor stages were pT2N0MX, pT2NXMX, and pT1NXMX, respectively. Findings by immunohistochemistry were similar but not identical for the three cases; markers for both proximal and distal parts of the nephron were expressed in each tumor, a finding that is in agreement with data from previous studies. The Ki-67-labeling index was below 5 in all three cases. Two of the tumors were predominantly hypodiploid (DNA-indexes 0.77 and 0.80), whereas the third tumor was hypertriploid (1.57) as measured by DNA-image cytometry. From the latter tumor live cells were available making it possible to establish its karyotype: 62-70,XXX,+del(X)(q11),-1,+2,+4,-5,-6,+7,-8,-9,-10,-11,+12,-13,-14,-15,+16,+17,+18,-19,+20,+21,-22[cp15]. Interphase fluorescence in situ hybridization analyses with centromere-specific probes for chromosomes 1, 3, 4, 6, 7, 9, 10, 17, 18, 20, and X showed that the two hypodiploid tumors had disomic and monosomic chromosome populations, whereas the karyotyped, near-triploid tumor was dominated by trisomic chromosome populations. Comparative genomic hybridization analysis was normal for the karyotyped tumor but abnormal for the two others. We conclude that multiple numerical chromosome aberrations may be a feature of mucinous tubular and spindle cell carcinomas of the kidney, but beyond that no clear-cut karyotypic aberration pattern is so far discernible.

Simultaneous bilateral breast carcinomas: a category with frequent coexpression of HER-2 and ER-alpha, high Ki-67 and bcl-2, and low p53.

The aim of this study was to evaluate clinicopathological characteristics and immunophenotypes of simultaneous bilateral adenocarcinomas of the breast and their axillary metastases. Immunohistochemical analysis and in situ hybridization were performed using formalin-fixed/paraffin-embedded tissues. In total, 15 primary and 9 metastatic tumors from 8 patients were evaluated. The expression of estrogen receptor-alpha (ER-alpha), progesterone receptor (PR), Ki 67, p53, bcl-2, and bax were evaluated by immunohistochemistry. Her2 gene amplification was evaluated by chromogenic in situ hybridization (CISH). Four patients were younger that 40 years of age (mean 47 years). Six patients had pleomorphic lobular carcinoma in 1 breast. Four of these had invasive ductal carcinoma in the contralateral breast. One patient had atypical medullary carcinoma in both breasts and 1 patient had atypical medullary carcinoma in 1 breast and pleomorphic lobular carcinoma in the other. The phenotype of the primary tumors and corresponding metastatic tumors was similar for the expression of ER-alpha (p=0.001), PR (p=0.03), and HER-2 (p=0.018). While strong coexpression of HER-2 and ER-alpha is exceptional in hereditary breast carcinoma and sporadic breast carcinoma, 6/8 (75%) patients in this study had tumors with strong coexpression of HER-2 and ER-alpha. P53 protein expression was found in only 2/15 (13%) primary tumors, which is in contrast to BRCA1-related hereditary bilateral breast carcinomas, which often express p53 protein. Most of the patients presented with axillary metastases and had very aggressive course. Characteristically, the tumors showed high levels of expression of ER-alpha and Her2 amplification, were bcl-2 positive, and had high Ki-67 fraction. However, in patients with atypical medullary carcinoma there was no expression of ER-alpha or amplification of Her-2.

Real-time PCR quantification of c-erbB-2 gene is an alternative for FISH in the clinical management of breast carcinoma patients.

Evaluation of gene amplification and protein expression of the c-erbB-2/neu in breast carcinomas has been an important task in breast cancer management. Although immunohistochemistry is widely applied, fluorescence in situ hybridization (FISH) technology shows its advantage in discriminating tumors in an objective manner. More recently, development of LightCycler technology permits evaluation of gene amplification with a small volume of DNA run in a 20 microL glass capillary. In this study, a series of 87 breast carcinomas were chosen for evaluation of c-erbB-2/neu gene amplification detected by both LightCycler technology and FISH. Real-time polymerase chain reaction (PCR) was performed in LightCycler capillaries with 10 ng sample DNA. By using LightCycler Relative Quantification Software version 1 (LightCycler, Roche, Mannheim, Germany), the amount of c-erbB-2 DNA was calculated as a ratio of c-erbB-2/reference gene quantity in a sample, and then the ratio was divided by the ratio of c-erbB-2 gene/reference gene quantities of a calibrator DNA (a standard DNA provided in the kit), which was run with each sample reaction in parallel. Dual-color FISH was performed on sections of the formalin-fixed, paraffin-embedded tissue array samples using the DAKO HER2 FISH pharmDX kit (DAKO A/S, Glostrup, Danmark) according to the manufacturer's instructions. Furthermore, immunohistochemistry was performed in parallel, with both the NCL-CB11 and HercepTest antibodies. Both the FISH technology and the LightCycler-PCR identified a similar percentage of tumors with c-erbB-2 gene amplification in our present study, 16% (14/87) and 15% (13/87), respectively, whereas immunohistochemistry demonstrated 32% and 34% c-erbB-2 overexpression with the NCL-CB11 and HercepTest antibodies, respectively. In addition, FISH and PCR were highly correlated in detecting tumors mainly with 3+++ c-erbB-2 protein expression by immunohistochemistry, indicating that LightCycler real-time quantification of c-erbB-2 gene may be an alternative to FISH in breast cancer clinical application.

Pilomatrix carcinoma with lymph node metastases.

Pilomatrix carcinoma is a rare skin tumor with an origin from hair matrix cells. The tumor is locally aggressive with a great tendency for recurrence, but the metastatic potential is limited. A pilomatrix carcinoma in 76-year-old female with lymph node metastases is presented. In addition to classical histopathological criteria and DNA ploidy analysis, a broad panel of antibodies was used for evaluation of the metastatic potential. Both primary tumor and lymph node metastasis revealed extremely high proliferation and apoptotic rates. High constant expressions of CD44v6 and P-cadherin were also observed. In the metastasis, significant reduction of E-cadherin and beta-catenin was detected. The best approach for assessment of metastatic potential of pilomatrix carcinoma seems to be the complex evaluation of routine histological criteria like vessel invasion, mitotic index, apoptotic count, and new molecular markers of cell death and adhesion.