Ming-Chieh Lin

The biomarkers of human papillomavirus infection in tonsillar squamous cell carcinoma—molecular basis and predicting favorable outcome

Presence of human papillomavirus (HPV) in variable proportions in tonsillar squamous cell carcinoma tissues has been demonstrated by several worldwide studies. Some reports emphasized the significance of HPV in predicting a better prognosis, as well as ethnic differences between Chinese and Caucasians. In order to understand the biological role of HPV and find out clinically accessible methods to determine its prognostic significance in primary tonsillar squamous cell carcinoma, we collected 92 patients with primary tonsillar squamous cell carcinoma diagnosed or treated in National Taiwan University Hospital, for whom archival tumor tissue were available. Immunohistochemical stains of p16INK4A, high-risk HPV in situ hybridization, and nested polymerase chain reaction (PCR)-based genechips were performed to detect HPV infection and determine its genotype. Clinical data were compared with HPV infection detected by the different methods mentioned above. Real-time PCR was also performed on the HPV16-positive [HPV16(+)] lesions to understand viral integration status. The positive rates of nested PCR-based genechips, overexpression of p16INK4A, and high-risk HPV in situ hybridization were 75% (69/92), 53% (49/92), and 44% (40/92), respectively. Both overexpression of P16INK4A and high-risk HPV in situ hybridization positivity were associated with favorable prognoses (P=0.004 and 0.001, respectively) and also independent prognostic factors in multivariate analyses (P=0.01 and 0.01, respectively). The positivity of nested PCR-based genechips was not statistically significant. From our data, primary tonsillar squamous cell carcinoma with positive immunohistochemical stains of p16INK4A and/or high-risk HPV in situ hybridization is associated with a better outcome, and both methods may serve as clinically accessible markers.

Loss of ARID1A expression and its relationship with PI3K-Akt pathway alterations and ZNF217 amplification in ovarian clear cell carcinoma

AT-rich interactive domain 1A (ARID1A) is a subunit of switch/sucrose non-fermentable (SWI/SNF) complex. Recently, alterations of ARID1A gene, phosphatidylinositol 3-kinase-protein kinase B (PI3K-Akt) pathway and zinc-finger protein 217 (ZNF217) gene have been identified as frequent molecular genetic changes in ovarian clear cell carcinoma. The relationships between these events have not been studied and integrated in the same cohort. This study was aimed at determining the correlation between these molecular events and other clinicopathological factors, including the prognostic impacts of these clinicopathological factors. A total of 68 ovarian clear cell carcinoma cases were collected and subjected to immunohistochemistry testing for ARID1A, SMARCA2, SMARCA4, SMARCB1 and phosphatase and tensin homolog (PTEN), mutation analysis for phosphatidylinositol-4,5-bisphosphate 3-kinase, catalytic subunit alpha (PIK3CA) gene and fluorescence in situ hybridization for ZNF217 amplification. The correlations between ARID1A expression, PI3K-Akt pathway, ZNF217 amplification and other clinicopathological factors were analyzed. Loss of ARID1A expression was present in 35 cases (52%) and loss of SMARCA2 expression occurred in 1 case. SMARCA4 and SMARCB1 expressions were preserved in all cases. PIK3CA mutations were present in 23 cases (34%) and loss of PTEN expression occurred in 8 cases (12%). Alterations in the PI3K-Akt pathway (PIK3CA mutations or loss of PTEN expression) were found in 42 cases (62%). ZNF217 amplification was detected in 21 cases (31%). Loss of ARID1A expression was significantly related to younger patient age (P=0.048), PI3K-Akt pathway activation (P=0.046) and ZNF217 amplification (P=0.028). All of the clinicopathological factors were not prognostic factors for ovarian clear cell carcinoma after multivariate analysis, except International Federation of Gynecology and Obstetrics staging (P=0.001). Our results showed that loss of ARID1A expression usually coexisted with PI3K-Akt pathway activation and/or ZNF217 amplification. Synergic effects of loss of ARID1A and PI3K-Akt pathway activation as well as ZNF217 amplification may be related to the development of ovarian clear cell carcinoma.

Epithelioid trophoblastic tumor of the broad ligament: a case report and review of the literature.

We report an epithelioid trophoblastic tumor, a recently delineated type of gestational trophoblastic tumor (GTT), discovered in the right broad ligament of a 41-year-old woman. The patient had gestational trophoblastic disease and was treated with methotrexate regimen 15 times 10 years earlier. The yellowish, spongy tumor with tiny hemorrhage spots was located in the right broad ligament, adherent to the right ovary. Microscopically, the tumor was circumscribed, with a pushing border, and the epithelial-differentiated tumor cells grew in cords, nests, and sheets within which were aggregates of hyaline material and necrotic debris. Most tumor cells were mononuclear with distinct cell borders, eosinophilic cytoplasm, and had nuclei with occasional indistinct nucleoli. Scattered multinucleated cells consistent with syncytiotrophoblastic cells were also present. Immunohistochemical staining revealed strong diffuse reactivity for cytokeratins (AE1/AE3, CAM 5.2, CK18), and focal reactivity, mainly in syncytiotrophoblastic cells, for beta-human chorionic gonadotropin, human placental lactogen, and inhibin-alpha. The histologic and immunohistochemical features were characteristic of epithelioid trophoblastic tumor and helped to distinguish the tumor from other trophoblastic tumors and squamous cell carcinoma. Our unusual findings in this case included a high level of serum beta-human chorionic gonadotropin, an unusual tumor location, and a higher Ki-67 proliferative index of 47.2%.

Nationwide Surveillance in Uterine Cancer: Survival Analysis and the Importance of Birth Cohort: 30-Year Population-Based Registry in Taiwan

Introduction Uterine cancer was the most rapidly increasing malignancy and the second most common gynecologic malignancy in Taiwan. Methods We analyzed the secular trend of uterine cancer incidence and compare the survival of women with uterine carcinomas and uterine sarcomas in Taiwan. Data on women diagnosed with uterine cancer between 1979 and 2008 were obtained from the Taiwan cancer registry. Survival data were analyzed by using Kaplan-Meier and Cox proportional hazards regression methods. Results Records of 11,558 women with uterine carcinomas and 1,226 women with uterine sarcomas were analyzed. The age-adjusted incidence rate of endometrioid adenocarcinoma increased from 0.83 per 100,000 women per year between 1979 and 1983 to 7.50 per 100,000 women per year between 2004 and 2008. The 5-year survival rate of women with endometrioid adenocarcinoma (83.2%) was higher than that for women with clear cell carcinoma (58.3%), serous carcinoma (54.4%), and carcinosarcoma (35.2%) (p<0.0001, log-rank test). The 5-year survival rates of women with low grade endometrial stromal sarcoma, endometrial stromal sarcoma, leiomyosarcoma (LMS), and adenosarcoma were 97.5%, 73.5%, 60.1%, and 77.2%, respectively (p<0.0001, log rank test). The histologic type of endometrioid adenocarcinoma, young age, and treatment period after 2000 were independent, favorable prognostic factors in women with uterine carcinomas by multivariate analysis. The histologic type of LMS, old age, and treatment period after 2000 were independent, poor prognostic factors in women with uterine sarcomas by multivariate analysis. Conclusions An increase over time in the number of patients with endometrioid adenocarcinomas was noted in this 30-year, nationwide, population-based study. Histologic type, age and treatment period were survival factors for uterine cancers. A more comprehensive assessment of uterine cancers and patient care should be undertaken on this increasingly common type of cancer.

Quantitative three-dimensional power Doppler ultrasound predicts the outcome of placental chorioangioma

The relationship of large and vascularized chorioangiomas to adverse pregnancy outcome is well recognized. We present a patient with a large placental tumor and signs of impending fetal cardiac failure. The angioarchitecture of the tumor depicted by three‐dimensional (3D) power Doppler ultrasound enabled us to accurately diagnose a placental chorioangioma. During the follow‐up period, quantitative flow data obtained using 3D power Doppler indicated altered hemodynamics in the tumor and concomitant improvement in the condition of the fetus, enabling us to manage the mother conservatively. Spontaneous delivery occurred at 38 weeks without any complications. This report demonstrates the potential value of 3D power Doppler in prenatal diagnosis and monitoring of pregnancies complicated by large, vascularized chorioangioma. Copyright

Increased Ki-67 proliferative index and absence of P16INK4 in CIN-HPV related pathogenic pathways different from cervical squamous intraepithelial lesion

Background/aim: It is generally assumed that similar pathways are involved in human papillomavirus (HPV) induced pathogenesis of cervical squamous intraepithelial lesions (SILs) and cancers and a subset of conjunctival intraepithelial neoplasm (CIN)—that the malignancies or pre-cancerous lesions arise through HPV oncoproteins E6 and E7, which disrupt the pathways of p53 and the product of the retinoblastoma (Rb) gene and, in turn, increase the protein product of gene p16INK4 through the mechanism of positive feedback. Several cell cycle molecules are detected to test this hypothesis. Methods: Nine cases of CIN and eight non-CIN cases were analysed for the expression of Ki-67, pRb, p53, and p16INK4 via immunohistochemistry. Nine cases of cervical high grade squamous intraepithelial lesion (HSIL), and 10 cases of cervical low grade squamous intraepithelial lesion (LSIL) were included for stain control of p16INK4a, and comparison of p16INK4a expression to CIN cases. A nested polymerase chain reaction and a genechip HPV typing were used to detect HPV infection and types in the CIN and non-CIN samples Results: HPV positivity was demonstrated in all of the CIN lesions but in none of the non-CIN lesions. The Ki-67 proliferative index (Ki-67 PI) was statistically higher in the CIN group than the non-CIN group; however, there were no differences of expression of pRb and p53 between the two groups and no expression of p16INK4 in all cases. All nine cases of HSIL, and seven out of 10 cases of LSIL used for stain control were immunoreactive for p16INK4a. There were statistically significant differences in overexpression of p16INK4a between the CINs and SILs Conclusions: The Ki-67 proliferative index may be a sensitive marker for CIN lesions and these results, with significant differences in overexpression of p16INK4a between CINs and SILs, may provide new evidence that HPV related mucosal dysplasia in different anatomical locations may lead to dissimilar molecular pathways.

Molecular alterations in endometrial and ovarian clear cell carcinomas: clinical impacts of telomerase reverse transcriptase promoter mutation.

Recently, mutations of telomerase reverse transcriptase (TERT) promoter were found in several types of cancer. A few reports demonstrate TERT promoter mutations in ovarian clear cell carcinomas but endometrial clear cell carcinoma has not been studied. The aims of this study were to compare differences of molecular alterations and clinical factors, and identify their prognostic impact in endometrial and ovarian clear cell carcinomas. We evaluated mutations of the TERT promoter and PIK3CA, expression of ARID1A, and other clinicopathological factors in 56 ovarian and 14 endometrial clear cell carcinomas. We found that TERT promoter mutations were present in 21% (3/14) of endometrial clear cell carcinomas and 16% (9/56) of ovarian clear cell carcinomas. Compared with ovarian clear cell carcinomas, endometrial clear cell carcinomas showed older mean patient age (P<0.001), preserved ARID1A immunoreactivity (P=0.017) and infrequent PIK3CA mutation (P=0.025). In ovarian clear cell carcinomas, TERT promoter mutations were correlated with patient age >45 (P=0.045) and preserved ARID1A expression (P=0.003). In cases of endometrial clear cell carcinoma, TERT promoter mutations were not statistically associated with any other clinicopathological factors. In ovarian clear cell carcinoma patients with early FIGO stage (stages I and II), TERT promoter mutation was an independent prognostic factor and correlated with a shorter disease-free survival and overall survival (P=0.015 and 0.009, respectively). In recurrent ovarian clear cell carcinoma patients with early FIGO stage, TERT promoter mutations were associated with early relapse within 6 months (P=0.018). We concluded that TERT promoter mutations were present in endometrial and ovarian clear cell carcinomas. Distinct molecular alteration patterns in endometrial and ovarian clear cell carcinomas implied different processes of tumorigenesis in these morphologically similar tumors. In ovarian clear cell carcinoma of early FIGO stage, patients with TERT promoter mutation require close follow-up during the initial 6 months following chemotherapy.

Downregulation of Brg-1 Repressed Expression of Cd44s in Cervical Neuroendocrine Carcinoma and Adenocarcinoma

Neuroendocrine carcinomas of the uterine cervix are rare tumors with early metastases, highly aggressive clinical behavior, and poor clinical outcome. Several adhesion molecules like cadherins have been tested in an attempt to explain their unique characteristics. Cluster differentiation 44 (CD44) is a widely expressed cell surface glycoprotein that serves as an adhesion molecule in cell-to-substrate and cell-to-cell interaction. We have examined the expression of the standard CD44 (CD44s) by immunohistochemical stains in the paraffin-embedded cervical neoplasm tissue of 17 cases of primary cervical neuroendocrine carcinoma, 28 cases of cervical adenocarcinoma, and 50 cases of cervical squamous cell carcinoma. Loss of CD44s expression was found in 16 of 17 neuroendocrine carcinomas, 14 of 28 adenocarcinomas, and three of 50 squamous cell carcinomas. The differences were statistically significant. We also examined immunohistochemically the expression of the BRG-1 subunit of the SWI-SNF complex, which has been reported to regulate the expression of CD44 in all cases. Loss of BRG-1 expression was observed in 12/16, 6/14, and 1/3 CD44s-negative neuroendocrine carcinomas, adenocarcinomas, and squamous cell carcinomas, respectively. This study suggests that loss of the CD44s molecule may imply special biological behaviors of cervical neuroendocrine carcinomas, and loss of expression of BRG-1 may contribute to this.

Hypermethylation of the CDKN2A gene promoter is a frequent epigenetic change in periocular sebaceous carcinoma and is associated with younger patient age

Periocular sebaceous carcinoma is an aggressive neoplasm with significant morbidity and mortality. Its pathogenesis is poorly understood. It is only rarely associated with Muir-Torre syndrome. Previous studies from Asian countries, have suggested that human papillomavirus (HPV) infection plays a role in the pathogenesis and overexpression of p16(INK4a), a surrogate marker of HPV infection, have also been reported. However, data from western countries seem contradictory. In order to clarify and explore the molecular and epigenetic basis of HPV, CDKN2A status and role of microsatellite instability in the development of periocular sebaceous carcinoma, 24 cases of periocular sebaceous carcinoma were analyzed for the expression of p16(INK4a) and mismatch repair proteins (MLH1, MSH2, MSH6 and PMS2) via immunohistochemistry. Nested polymerase chain reaction (PCR) and genechip HPV typing were used to detect HPV infection and decide its genotype when present. PCR amplification using a consensus primer pair was also performed to detect β-HPV. The methylation status of CDKN2A promoter region was studied by methylation-specific polymerase chain reaction. HPV-positivity was demonstrated in only one of our cases (HPV 16), while another case showed p16(INK4a) overexpression. All cases showed preserved expression of mismatch repair proteins. CDKN2A promoter hypermethylation was noted in nearly half of our cases (11/24) and was associated with younger patient age (P = .013). Our results showed that periocular sebaceous carcinoma is rarely associated with HPV and microsatellite instability. Higher frequency of CDKN2A promoter hypermethylation in younger patients implies a significant epigenetic role in tumor development in this age group.

Concurrent Endometrial Carcinoma in Patients with a Curettage Diagnosis of Endometrial Hyperplasia

BACKGROUND/PURPOSE Endometrial hyperplasia is considered a precursor of endometrial carcinoma, but concurrent endometrial carcinoma in patients with endometrial hyperplasia is seen frequently. Our aim was to examine the risk factors for coexisting endometrial carcinoma in patients with endometrial hyperplasia. METHODS Between January 1996 and September 2006, 77 patients who underwent hysterectomy for endometrial hyperplasia were enrolled retrospectively. We divided the patients into non-endometrial carcinoma and endometrial carcinoma groups, depending on the final pathology of hysterectomy and analyzed the clinical variables of these patients. RESULTS The prevalence rate of concurrent endometrial carcinoma in patients with endometrial hyperplasia was 26%. Those with atypical endometrial hyperplasia had a higher rate of coexisting endometrial carcinoma (54%). In addition to cytologic atypia, body mass index (BMI) was another risk factor. All the patients with concomitant endometrial carcinoma had at least one risk factor, but almost 50% of the cases in the non-endometrial group had no risk factors. Half of the women with cytological atypia and BMI > 25 had coexisting endometrial carcinoma. CONCLUSION When patients are diagnosed with endometrial hyperplasia, surgical intervention should be performed in those with cytological atypia and higher BMI because of the possibility of coexisting endometrial carcinoma.