Immunohistochemistry in Kaposi's sarcoma.

The importance of immunohistochemistry (IHC) to our understanding, ability to confidently diagnose and treat Kaposi's sarcoma (KS) has grown steadily in the past few decades. IHC has been performed on many KS specimen types, with > 100 different primary antibodies. Therefore, it is not surprising that IHC has helped unravel the histogenesis, understand the pathogenesis and facilitate the diagnosis of KS and identify novel therapeutic targets in the disease. This paper reviews the literature on the use of IHC in the study of KS.

PLD1 is overexpressed in an ER-negative MCF-7 cell line variant and a subset of phospho-Akt-negative breast carcinomas.

We have used a novel variant of the human oestrogen receptor (ER)-positive MCF-7 cell line, TMX2-28, as a model to study breast cancer. TMX2-28 cells show no detectable levels of mRNA or protein expression for the ER and express basal cytokeratins (CKs) 5, 14, and 17. cDNA microarray comparison between TMX2-28 and its parent cell line, MCF-7, identified 1402 differentially expressed transcripts, one of which was, phospholipase D1 (PLD1). Using real-time RT-PCR, we confirmed that PLD1 mRNA levels are 10-fold higher in TMX2-28 cells than in MCF-7 cells. We next examined PLD1 expression in human breast carcinomas. Phospholipase D1 mRNA levels were higher in breast tumours that expressed high-mRNA levels of basal CKs 5 and/or 17, but PLD1 mRNA levels were not significantly higher in ER-negative tumours. Phospholipase D1 protein was overexpressed in 10 of 42 (24%) breast tumours examined by IHC. Phospholipase D1 was overexpressed in 6 of 31 ER-positive tumours and 4 of 11 ER-negative tumours. Phospholipase D1 was overexpressed in three of the four tumours that showed high CK5/17 expression. Five PLD1-positive tumours were negative for phospho-Akt expression, but positive for phospho-mammalian target of rapamycin (mTOR) expression. The other five PLD1-positive breast tumours showed positive expression for phospho-Akt; however, only two of these cases were positive for phospho-mTOR. In this study, we report that PLD1 and phospho-mTOR are coexpressed in a subset of phospho-Akt-negative breast carcinomas.

Combined carcinoid tumor and ganglioneuroblastoma of the lung: a case report.

Carcinoid tumor is a well-differentiated epithelial neuroendocrine neoplasm which is common in the lung. Ganglioneuroblastoma is a moderately differentiated nonepithelial neuroendocrine neoplasm which is very rare in the lung. Neuroendocrine tumors with epithelial and nonepithelial elements are rare in any site and have not been reported in the lung. This case is an example of a primary neuroendocrine lung tumor combining epithelial and non-epithelial components: carcinoid tumor and ganglioneuroblastoma.

Tyrosine kinase activation in breast carcinoma with correlation to HER-2/neu gene amplification and receptor overexpression.

The HER-2/neu oncogene encodes a transmembrane receptor with intrinsic tyrosine kinase activity. A pilot study was performed to investigate downstream effects of HER-2/neu (or related growth factor receptor) activation by identifying phosphorylated tyrosine. Fifty-four breast carcinomas were evaluated for HER-2/neu overexpression by the HercepTest (Dako, Carpinteria, CA) and the monoclonal CB11 antibody (Ventana, Tucson, AZ). Phosphotyrosine (an indication of tyrosine kinase activity) was detected by an antiphosphotyrosine mouse monoclonal antibody (Upstate Biotechnology, Lake Placid, NY). The gene amplification status was evaluated in 50 of the 54 cases by fluorescence in situ hybridization (FISH) using the Ventana gene probe. The HER-2/neu oncogene amplification was detected in 28% (14 of 50) of cases. Of the 14 cases showing oncogene amplification, tyrosine kinase activity was detected in 9 (64.2%) cases. There was moderate agreement between HER-2/neu gene amplification and tyrosine kinase activity (kappa = 0.43). Immunohistochemical staining of 3+ (with both HercepTest and CB11) showed better agreement with HER-2/neu oncogene amplification and increased tyrosine kinase activity than 2+ immunohistochemical staining. Overall, oncogene amplification and overexpression correlated with increased tyrosine kinase activity, supporting the mechanism of tyrosine kinase activation by HER-2/neu amplification and overexpression. However, 7 cases showing increased tyrosine kinase activity did not show gene amplification or 3+ receptor expression (by either HercepTest or CB11), raising the possibility of other growth factor receptors operating via the tyrosine kinase pathway. There was no apparent correlation between tyrosine kinase activity and hormone receptor status (estrogen or progesterone). Increased tyrosine kinase activity is more commonly associated with higher-grade tumors and thus may correlate with aggressive biologic behavior in breast carcinoma. The results of this pilot study suggest that a larger-scale investigation into downstream activation of tyrosine kinase and correlation to clinical outcome or response to Herceptin therapy may identify subsets of patients whose clinical response or outcome may be predicted by tyrosine kinase activation.

Pleomorphic lobular carcinoma: morphology, immunohistochemistry, and molecular analysis.

Infiltrating pleomorphic lobular carcinoma (PLC) is an aggressive variant of infiltrating lobular carcinoma. Recently, in situ changes identical to PLC (PLCIS) have been described. The role of prognostic markers and their correlation with therapeutics, clinical outcome, and genetic changes is not well established in PLC. The authors examined 38 cases of this entity to understand better this tumor's biology. Immunohistochemical (IHC) analysis was performed in 21 specimens for estrogen and progesterone steroid receptors, p53, Her 2 (p185), and GCDFP-15. Genomic deoxyribonucleic acid was obtained from microdissected tumor as well as normal control cells, and loss of heterozygosity was investigated at the ESR (16q24), p53 (TP53 17p), Her 2 (17q 11-12), and BRCA 1 (17q12-25) loci. In this series, the average patient age was 57.5 years (age range, 24-92 years). Twenty-seven women were postmenopausal. Tumor size ranged from 1.2 to 25 cm. Six patients were a pathologic stage I; 19, stage II; 12, stage III; and one, stage IV. Histologically, multifocal nodular aggregates of discohesive pleomorphic tumor cells were seen interspersed in dense and fibrotic breast parenchyma. Twenty-nine percent of the specimens demonstrated associated signet ring cells. The remainder had dishesive, globoid, plasmacytoid cells with high-grade nuclear features. PLCIS was identified in 17 of 38 patients (45%), and lobular carcinoma in situ (LCIS) was noted in 8 patients (21%). IHC analysis showed estrogen immunoreactivity in 81%, progesterone in 67%, GCDFP-15 in 71%, and Her 2 in 81% (2+ to 3+ membranous staining) of specimens. Antibodies to p53 stained the tumor cell nuclei in 48% of the tumors. Loss of heterozygosity was identified in 52% of the specimens at the p53 locus, 18% at the ESR locus, 19% to 24% at the Her 2 loci, and 27% to 32% at the BRCA 1 locus. Follow-up was available in 19 patients and ranged from 12 months to 15 years (mean, 73 months). Seven patients had no evidence of disease at last examination (range, 1-15 years), three patients were alive with disease (range, 2-14 years), and nine patients were dead of disease (range, 2 months-9 years). Six patients had subsequent diagnoses of tumor in the contralateral breast. Analysis shows that PLC tends to appear in older postmenopausal women who present with locally advanced disease. PLCIS was found to be associated with PLC 45% of the time. The aggressive clinical course of patients with PLC is supported by tumor immunoreactivity with unfavorable markers Her 2 and p53. Overexpression of Her 2 in PLC may be therapeutically relevant, enabling the use of novel chemotherapeutic drugs like Herceptin. Interestingly, tumors that were Her 2 immunoreactive also maintained estrogen hormone immunoreactivity.

Loss of heterozygosity in P53, BRCA1, and estrogen receptor genes and correlation to expression of p53 protein in ovarian epithelial tumors of different cell types and biological behavior.

Loss of heterozygosity (LOH) of tumor suppressor genes (TSGs) in ovarian epithelial tumors of differing cell types and biological behavior has not been thoroughly investigated. Moreover, there have been conflicting reports correlating LOH of the p53 gene to overexpression of p53 protein. This study evaluated 34 formalin-fixed, paraffin-embedded ovarian epithelial tumors for LOH by polymerase chain reaction (PCR) for the following microsatellite markers: TP53(17p13.1/p53 gene), D17S579(17q/BRCA1 gene), and ESR (6q24-27/estrogen receptor gene). LOH of the TP53 marker was detected in 4 (44%) of 9 informative serous cystadenocarcinomas (SCa) but in 0 of 4 informative clear cell carcinomas (CCa) and 0 of 5 informative serous tumors of low malignant potential (SLMP). LOH of the BRCA1 marker was detected in 5 (83%) of 6 informative SCa, but in 1 (13%) of 8 informative CCa and 1 (14%) of 7 informative SLMP. LOH of the ESR marker was detected in 4 (50%) of 8 informative SCa, but in 0 of 4 informative CCa and 1 (16%) of 6 informative SLMP. p53 protein overexpression was present in 8 of 12 SCa but did not correlate to TP53 LOH. LOH for TP53, D17S579/ BRCA1, and ESR is common in ovarian SCa, and is observed in primary tumors as well as metastases. In contrast, these genetic alterations are less common in CCa and in the biologically less aggressive SLMP tumors. These data suggest different mechanisms of oncogenesis in ovarian epithelial tumors of different cell types and biological behavior.

The unsatisfactory margin in breast cancer surgery.

BACKGROUND: Surgical margin involvement with breast cancer usually results in obligatory reexcision or mastectomy. While unalterable occult host and pathologic factors may interfere with margin clearance during the initial excision, it is possible that alterations in surgical technique might increase the likelihood of obtaining satisfactory margins. METHODS: Two hundred and thirty-five patients who were candidates for breast conservation therapy were identified for 1991 and 1996 using the Tumor Registry. Margins were defined as "unsatisfactory" if there was microscopic involvement with tumor or the margin was close at initial excisional biopsy and the surgeon opted for reexcision. Multiple logistic regression analyses of factors associated with margin status were performed. RESULTS: One hundred thirty-two (56%) patients had positive or close (unsatisfactory) margins; this rate increased from 51% in 1991 to 59% in 1996. Patients with unsatisfactory margins underwent more procedures (mean 2.0 versus 1.2; P <0.0001) than patients whose margins were satisfactory. The breast conservation rate for patients with unsatisfactory margins was 64% compared with 99% for patients with satisfactory margins. A multiple logistic regression demonstrated that patients with unsatisfactory margins were 67 times more likely to have a mastectomy than patients whose margins were satisfactory after adjusting for other significant factors (P <0.0001). The practice of fine needle aspiration biopsy, orientation of specimen margins by the surgeon, and reexcision of tumor at the first operation were statistically significant technical factors in obtaining satisfactory margins. Significant pathology factors were extensive intraductal component (EIC), lobular or ductal extension, and tumor size. CONCLUSION: These data show that technical factors in the surgical management of breast cancer, as well as biological factors such as EIC, can influence the success of breast conservation.

The use of microsatellite instability in the distinction between synchronous endometrial and colonic adenocarcinomas.

The association of endometrial carcinoma with other gynecologic neoplasms, especially ovarian and fallopian tube carcinoma, has been well documented and is usually interpreted as a result of a field defect. Sporadic synchronous primary carcinomas occurring in the endometrium and colon are extremely rare, especially in the absence of the familial genetic abnormalities seen in hereditary nonpolyposis colorectal carcinoma (HNPCC) syndrome, and may present a diagnostic dilemma. Two cases of synchronous adenocarcinomas of the endometrium and colon were studied for genetic abnormalities and differences to test for the presence of two primary tumors. Primary tumors, metastases, and normal tissues were microdissected from formalin-fixed, paraffin-embedded tissues. PCR amplification was performed for microsatellite DNA markers on chromosome 17q and 11q13. The colonic tumors were moderately and poorly differentiated, invasive, nonmucinous adenocarcinomas, whereas one uterine tumor was endometrioid adenocarcinoma and the other was papillary serous carcinoma. Although microsatellite instability, as evidenced by changes in the lengths of the amplified PCR products, was detected at 17q and 11q13 loci in the uterine and colonic neoplasms, the patterns of instability differed between the two primary tumor sites. Moreover, the lymph node metastasis in one colonic tumor had genetic alterations that differed from that of the primary tumor. In both patients, the molecular studies suggested the presence of two synchronous primary tumors. Molecular techniques may assist in distinguishing two separate primaries by determining the contraction and expansion of microsatellite regions in DNA obtained by microdissection from the primary tumors and associated metastases.

Management of advanced juvenile granulosa cell tumor of the ovary.

Juvenile granulosa cell tumors (JGCTs) of the ovary are rare. They usually present in children and adolescents. About 90% are diagnosed in early stage (FIGO I) with a favorable prognosis. More advanced stages (FIGO II-IV) have a poor clinical outcome. We report two cases of short-term, disease-free survival of teenagers with Stage III JGCTs treated with aggressive debulking and thorough staging but conservative surgery relative to the uterus and contralateral tube and ovary plus carboplatin and etoposide chemotherapy. These results are encouraging, but the best treatment for extensive and recurrent disease has yet to be determined.

Uterine adenomatoid tumors: immunohistochemical characteristics with emphasis on Ber-EP4 immunoreactivity and distinction from adenocarcinoma.

Uterine adenomatoid tumors (UATs) may be difficult to distinguish from metastatic adenocarcinoma, particularly in pelviscopic biopsy specimens, myomectomy specimens, or endometrial samplings. This problem may arise because of the infiltration of vacuolated mesothelial cells between fascicles of prominent smooth muscle, which is a characteristic feature in UAT. This diagnostic difficulty has led to inappropriate surgery as reported in the past. Eleven UATs were studied to clarify the differences between these tumors and metastatic adenocarcinoma. Six of the tumors were grossly indistinguishable from leiomyomas. Histologically, the neoplastic mesothelial cells diffusely infiltrated smooth muscle fascicles in all cases, mimicking the pattern of metastatic adenocarcinoma. Immunohistochemical studies using antikeratin and Ber-EP4 antibodies were positive in the mesothelial component in all 11 and nine tumors, respectively, whereas all 11 UATs were negative using antibodies to carcinoembryonic antigen, CD15, TAG-72, and epithelial membrane antigen. Immunoreactivity using Ber-EP4 was focal, membranous, and usually weak, although a strong signal was present in one case. Immunoreactivity using Ber-EP4 (compared with negativity reported for mesothelial proliferations of other sites) may be related to the unique müllerian characteristic of the female peritoneum. Although nonimmunoreactivity for Ber-EP4 favors a diagnosis of UAT over that of adenocarcinoma, Ber-EP4 immunoreactivity does not exclude UAT.

Reliability of quantitating proliferative cell nuclear antigen in epithelial ovarian tumors.

Sources of variability in quantitating proliferative cell nuclear antigen (PCNA) by image analysis were evaluated in paraffin sections of 18 ovarian tumors of low malignant potential (LMP) and grade 1 (G1) carcinomas. The correlation coefficient of reliability (R) was calculated to determine how reliable a single observation was for representing a "true" tumor value. Reliability of 61% was obtained when interobserver and intraobserver variability were assessed. Threshold settings for positive nuclear and antibody signals minimally affected the overall reliability. The reliability of a single block of tumor for representing true tumor value was R = 0.61. These sources of variability render this technique impractical for evaluating proliferative characteristics in LMP and G1 common epithelial ovarian tumors with PCNA staining.

Nd:YAG laser excision of a giant gingival pyogenic granuloma of pregnancy.

A 19-year-old Hispanic nullipara experienced the rapid growth of an oral lesion on the right lower gingiva which she had first noticed at 29 weeks gestation. The lesion interfered with oral hygiene and eating. At surgery, the lesion measured 3.5 x 2.5 x 2.0 cm. Biopsy confirmed a pyogenic granuloma ("granuloma gravidarum"). Panorex films showed no bony invasion. The lesion was excised using the Nd:YAG laser under general anesthesia when the patient had reached 36 3/7 weeks gestation. We chose the Nd:YAG laser over the CO2 laser for the removal of this very vascular lesion, because of its superior coagulation characteristics. This technique results in removal of the lesion with less danger of bleeding than with any other surgical technique. The patient did well postoperatively, delivered a healthy 3,884 g infant at 40 6/7 weeks gestation, and has had no recurrence after 15 months of follow-up.

Primary breast cancer of the vulva.

Supernumerary breasts on the vulva are a rare occurrence. There have been 26 cases previously reported and only 4 case reports of primary mammary carcinoma of the vulva. This is the fifth case report of mammary carcinoma of the vulva and the second report using tamoxifen for adjuvant treatment.

Cushing's syndrome secondary to malignant lipoid cell tumor of the ovary.

Malignant lipoid cell tumors of the ovary are rare lesions that are frequently associated with endocrinologic abnormalities. A case of a woman with this lesion who developed Cushing's syndrome with progression of tumor is presented. Neither aggressive medical therapy with ketoconazole nor multiagent chemotherapy was beneficial in controlling tumor growth or physical and biochemical manifestations of Cushing's syndrome.

Management of advanced juvenile granulosa cell tumor of the ovary.

Juvenile granulosa cell tumor of the ovary occurs most frequently in young women and children. Ten percent of cases present during pregnancy. The majority of tumors are in FIGO Stage I and have a favorable prognosis. The prognosis of higher stage tumors, however, is generally less favorable. We report the long-term, disease-free survival of a patient with FIGO Stage III juvenile granulosa cell tumor of the ovary. We believe this to be the first report of a successful pregnancy following "MAC" chemotherapy for this particular malignancy.

Intermediate filamentous proteins in adult granulosa cell tumors. An immunohistochemical study of 25 cases.

Adult granulosa cell tumors (AGCTs) are classified as sex cord-stromal tumors of the ovary. However, they may be confused with other primary ovarian neoplasms. Intermediate filaments, specifically vimentin and cytokeratins, have been identified in AGCTs by immunohistochemistry performed on frozen and formalin-fixed, paraffin-embedded tissue and two-dimensional electrophoresis. Recently, however, immunohistochemical demonstration of cytokeratin has been used as supporting evidence of epithelial rather than sex cord-stromal differentiation in ovarian neoplasia. To investigate further intermediate filamentous proteins in AGCTs, 25 such tumors were studied by immunohistochemistry in formalin-fixed, paraffin-embedded sections. Cytoplasmic staining was observed, frequently in a distinct punctate, paranuclear pattern, in 14 of 25, 14 of 25, and seven of 17 tumors using monoclonal antibodies AE1/AE3, CAM 5.2, and 35BH11, respectively, which share the ability to detect low molecular weight cytokeratins. Staining for cytokeratin was not seen in any of the 17 tumors studied using the antibody 34BE12. Twenty-three of 25 tumors showed strong positivity for vimentin, characteristically seen as globoid paranuclear staining. Nine of 25 tumors contained desmin, which was restricted to the intermixed spindle cell, cortical type stromal component of the tumors. These patterns of immunoreactivity for intermediate filaments, particularly cytokeratins, are different than in common epithelial tumors of the ovary and may be useful in the differential diagnosis of ovarian neoplasia. Moreover, the immunohistochemical detection of cytokeratins should not be used as a criterion for excluding AGCT from the differential diagnosis of an ovarian neoplasm.

Ovarian serous borderline epithelial tumors with multiple retroperitoneal nodal involvement: metastasis or malignant transformation of epithelial glandular inclusions?

One of four patients who underwent lymph node excision at exploration for ovarian serous borderline epithelial tumor (OSBT) at Baystate Medical Center was found to have FIGO Stage III C lesion associated with extensive ovarian external (surface) papillary growth, peritoneal implants in the omentum and cul-de-sac, and involvement of multiple pelvic and periaortic lymph nodes by the tumor. Histologically, the lymph nodes showed an admixture of endosalpingeal glandular inclusions with neoplastic tissue identical to the ovarian tumor. The exact histogenesis and the prognostic significance of the nodal involvement by OSBT are still not fully understood. Although there is a small number of reported cases of lymph node involvement associated with OSBT, they are described as examples of nodal metastases or independent primary foci of malignant transformation. This paper presents an interesting association of OSBT with extensive pelvic and periaortic nodal involvement and reviews the relevant literature.

Histiocytosis X of the vulva: a case report and review of the literature.

Histiocytosis X of the female genital tract is unusual. Thirty-two cases have been reported to date in the world literature. An additional case is reported herein, presenting as a vulvar ulcer in a 2.5-year-old child with osteolytic lesions of the skull, splenomegaly, and otitis media. The diagnosis of histiocytosis X may be established by identifying the Langerhans histiocyte, characterized by nuclear grooves, immunoreactivity for S-100 protein, and pentalamellar cytoplasmic structures seen by electron microscopy. Prognosis is difficult to determine with certainty. However, age of less than 2 years at presentation, multi-organ involvement, and/or organ dysfunction appear to be associated with a less favorable prognosis. The patient presented herein is currently receiving vinblastine chemotherapy for recurrence of disease, manifested as an osteolytic lesion in the skull.