Recommendations for pathologic practice using digital pathology: consensus report of the Korean Society of Pathologists

Digital pathology (DP) using whole slide imaging (WSI) is becoming a fundamental issue in pathology with recent advances and the rapid development of associated technologies. However, the available evidence on its diagnostic uses and practical advice for pathologists on implementing DP remains insufficient, particularly in light of the exponential growth of this industry. To inform DP implementation in Korea, we developed relevant and timely recommendations. We first performed a literature review of DP guidelines, recommendations, and position papers from major countries, as well as a review of relevant studies validating WSI. Based on that information, we prepared a draft. After several revisions, we released this draft to the public and the members of the Korean Society of Pathologists through our homepage and held an open forum for interested parties. Through that process, this final manuscript has been prepared. This recommendation contains an overview describing the background, objectives, scope of application, and basic terminology; guidelines and considerations for the hardware and software used in DP systems and the validation required for DP implementation; conclusions; and references and appendices, including literature on DP from major countries and WSI validation studies.

Recommendations for pathologic practice using digital pathology: consensus report of the Korean Society of Pathologists

Digital pathology (DP) using whole slide imaging (WSI) is becoming a fundamental issue in pathology with recent advances and the rapid development of associated technologies. However, the available evidence on its diagnostic uses and practical advice for pathologists on implementing DP remains insufficient, particularly in light of the exponential growth of this industry. To inform DP implementation in Korea, we developed relevant and timely recommendations. We first performed a literature review of DP guidelines, recommendations, and position papers from major countries, as well as a review of relevant studies validating WSI. Based on that information, we prepared a draft. After several revisions, we released this draft to the public and the members of the Korean Society of Pathologists through our homepage and held an open forum for interested parties. Through that process, this final manuscript has been prepared. This recommendation contains an overview describing the background, objectives, scope of application, and basic terminology; guidelines and considerations for the hardware and software used in DP systems and the validation required for DP implementation; conclusions; and references and appendices, including literature on DP from major countries and WSI validation studies.

Mutational signatures and chromosome alteration profiles of squamous cell carcinomas of the vulva

Vulvar squamous cell carcinoma (SCC) consists of two different etiologic categories: human papilloma virus (HPV)-associated (HPV (+)) and HPV-non-associated (HPV (−)). There have been no genome-wide studies on the genetic alterations of vulvar SCCs or on the differences between HPV (+) and HPV (−) vulvar SCCs. In this study, we performed whole-exome sequencing and copy number profiling of 6 HPV (+) and 9 HPV (−) vulvar SCCs and found known mutations (TP53, CDKN2A and HRAS) and copy number alterations (CNAs) (7p and 8q gains and 2q loss) in HPV (−) SCCs. In HPV (+), we found novel mutations in PIK3CA, BRCA2 and FBXW7 that had not been reported in vulvar SCCs. HPV (−) SCCs exhibited more mutational loads (numbers of nonsilent mutations and driver mutations) than HPV (+) SCCs, but the CNA loads and mutation signatures between HPV (+) and HPV (−) SCCs did not differ. Of note, 40% and 40% of the 15 vulvar SCCs harbored PIK3CA and FAT1 alterations, respectively. In addition, we found that the SCCs harbored kataegis (a localized hypermutation) in 2 HPV (+) SCCs and copy-neutral losses of heterozygosity in 4 (one HPV (+) and 3 HPV (−)) SCCs. Our data indicate that HPV (+) and HPV (−) vulvar SCCs may have different mutation and CNA profiles but that there are genomic features common to SCCs. Our data provide useful information for both HPV (+) and HPV (−) vulvar SCCs and may aid in the development of clinical treatment strategies.

Patient-Derived Xenograft Models of Epithelial Ovarian Cancer for Preclinical Studies / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: Patient-derived tumor xenografts (PDXs) can provide more reliable information about tumor biology than cell line models. We developed PDXs for epithelial ovarian cancer (EOC) that have histopathologic and genetic similarities to the primary patient tissues and evaluated their potential for use as a platform for translational EOC research. MATERIALS AND METHODS: We successfully established PDXs by subrenal capsule implantation of primary EOC tissues into female BALB/C-nude mice. The rate of successful PDX engraftment was 48.8% (22/45 cases). Hematoxylin and eosin staining and short tandem repeat analysis showed histopathological and genetic similarity between the PDX and primary patient tissues. RESULTS: Patients whose tumors were successfully engrafted in mice had significantly inferior overall survival when compared with those whose tumors failed to engraft (p=0.040). In preclinical tests of this model, we found that paclitaxel-carboplatin combination chemotherapy significantly deceased tumor weight in PDXs compared with the control treatment (p=0.013). Moreover, erlotinib treatment significantly decreased tumor weight in epidermal growth factor receptor–overexpressing PDX with clear cell histology (p=0.023). CONCLUSION: PDXs for EOC with histopathological and genetic stability can be efficiently developed by subrenal capsule implantation and have the potential to provide a promising platform for future translational research and precision medicine for EOC.

Extrapulmonary Lymphangioleiomyoma: Clinicopathological Analysis of 4 Cases

BACKGROUND: Lymphangioleiomyomatosis (LAM) is a slowly progressive neoplastic disease that predominantly affects females. Usually, LAM affects the lung; it can also affect extrapulmonary sites, such as the mediastinum, the retroperitoneum, or the lymph nodes, although these locations are rare. A localized form of LAM can manifest as extrapulmonary lesions; this form is referred to as extrapulmonary lymphangioleiomyoma (E-LAM). Due to the rare occurrence of E-LAM and its variable, atypical location, E-LAM is often difficult to diagnose. Herein, we report the clinicopathological information from four E-LAM cases, and also review previous articles investigating this disease. METHODS: Four patients with E-LAM were identified at the Samsung Medical Center (Seoul, Korea) from 1995 to 2012. All E-LAM lesions underwent surgical excision. RESULTS: All patients were females within the age range of 43 to 47 years. Two patients had para-aortic retroperitoneal masses, while the other two patients had pelvic lesions; two out of the four patients also had accompanying pulmonary LAM. In addition, no patient displayed any evidence of tuberous sclerosis. Histologically, two patients exhibited nuclear atypism with cytologic degeneration. CONCLUSIONS: E-LAM should be considered in the differential diagnosis of patients presenting with pelvic or para-aortic masses. We also conclude that further clinical and pathological evaluation is needed in patients with E-LAM and nuclear atypism.

Synchronous gynecologic malignancy and preliminary results of Lynch syndrome / 부인종양

OBJECTIVE: Lynch syndrome is a hereditary cancer syndrome that increases the risks of colorectal and gynecologic malignancies such as endometrial and ovarian cancer. Studies have shown that mutations in mismatch repair genes (MSH2, MSH6, and MLH1) are associated with Lynch syndrome. The aim of our study was to estimate the value of MSH2, MSH6, and MLH1 immunohistochemistry based on family history in a Korean sample. METHODS: Thirty six women with synchronous gynecologic tumors of endometrial and ovarian cancer were identified among patients being treated at our institution. Among them, 32 patients had tumor blocks (total 62 slides) available for analysis. According to a diagnostic algorithm, we performed immunohistochemistry analyses. Staining was scored based on intensity and proportion (negative or 0: intensity undetectable or minimal, proportion <5%; weak or 1+: intensity mild, proportion 5-30%; strong or 2+: intensity moderate to marked, proportion 30-99%). RESULTS: Among 32 eligible patients, 9 (28%) had a family history of cancer. Six patients (19%) were negative for MLH1; among them, four (4/6) were negative at both sites. Nine patients (28%) were negative for MSH2 or MSH6 at both sites or negative for both MSH2 and MSH6. Among these three patients showed negative staining for both sites. The three patients showing negative staining for MLH1, MSH2, and MSH6 at both sites with family history were considered to be the screening positive groups of Lynch syndrome. CONCLUSION: In this study, the frequency of Lynch syndrome associated immunohistochemical staining (MLH1, MSH2, and MSH6) group was estimated as 9% (3/32) among Korean women with synchronous gynecologic tumors.

AGR2, a mucinous ovarian cancer marker, promotes cell proliferation and migration

Ovarian cancer is a leading cause of death in women. Early detection of ovarian cancer is essential to decrease mortality. However, the early diagnosis of ovarian cancer is difficult due to a lack of clinical symptoms and suitable molecular diagnostic markers. Thus, identification of meaningful tumor biomarkers with potential clinical application is clearly needed. To search for a biomarker for the early detection of ovarian cancer, we identified human anterior gradient 2 (AGR2) from our systematic analysis of paired normal and ovarian tumor tissue cDNA microarray. We noted a marked overexpression of AGR2 mRNA and protein in early stage mucinous ovarian tumors compared to normal ovarian tissues and serous type ovarian tumors by Western blot analysis and immunohistochemistry. To further elucidate the role of AGR2 in ovarian tumorigenesis, stable 2774 human ovarian cancer cell lines overexpressing AGR2 were established. Forced expression of AGR2 in 2774 cells enhanced the growth and migration of ovarian cancer cells. AGR2 protein was detected in the serum of mucinous ovarian cancer patients by Western blot and ELISA analysis. Thus, AGR2 is a potential biomarker for the diagnosis of mucinous ovarian cancer and an ELISA assay may facilitate the early detection of mucinous ovarian cancer using patient serum.

Papillary Thyroid Carcinoma of a Diffuse Sclerosing Variant: Ultrasonographic Monitoring from a Normal Thyroid Gland to Mass Formation

A diffuse sclerosing variant of papillary thyroid carcinoma is uncommon and has a tendency for rapid growth and a higher incidence of cervical lymph node metastases. We experienced a case of a diffuse sclerosing variant of papillary thyroid carcinoma in a 48-year-old man. This case showed benign features on initial ultrasonography and positron emission tomography (PET) scan. A new nodule was detected on follow-up ultrasonography that showed rapid enlargement. This case was confirmed by surgical excision. We herein describe the initial and follow-up ultrasonographic findings of a diffuse sclerosing variant of papillary thyroid carcinoma.

A Case of Congenital Hepatic Fibrosis in Kabuki Syndrome / 대한소아소화기영양학회지

Kabuki syndrome is characterized by peculiar facial features, developmental delay, and mental retardation. Congenital hepatic abnormalities in Kabuki syndrome patients have been sporadically reported in the literature and consist of extrahepatic biliary atresia, neonatal sclerosing cholangitis, and transient neonatal cholestasis. We report here a case of congenital hepatic fibrosis in a patient with Kabuki syndrome. To our knowledge, only one case of congenital hepatic fibrosis has been reported in the setting of Kabuki syndrome.

Ghrelin Levels in Gastric Mucosa before and after Eradication of Helicobacter pylori

BACKGROUND/AIMS: The relationship between Helicobacter pylori infection and ghrelin is controversial. We compared ghrelin levels in gastric mucosa and plasma between H. pylori-positive and -negative subjects, and between before and after H. pylori eradication. METHODS: We compared the ghrelin levels in the antrum, body, and fundus between H. pylori-positive and -negative subjects; in stomach tissues between before and after H. pylori eradication; and in plasma and tissue in 10-person cohorts between before and after H. pylori eradication therapy. Body mass index, age, and sex were controlled for when comparing ghrelin levels. RESULTS: Stomach ghrelin levels (in the antrum, body, and fundus) did not differ significantly between H. pylori-positive and -negative samples (p=0.095, 0.316, and 0.897, respectively), or between before and after H. pylori eradication (p=0.19, 0.178, and 0.513, respectively). In the ten-person cohort study, plasma ghrelin levels in the eight H. pylori-positive subjects were 2,260 pg/mL (range, 1,280-3,770 pg/mL) and 1,900 pg/mL (range, 1,350-5,200 pg/mL) before and after eradication therapy (p=0.871). Stomach ghrelin levels did not differ significantly in the eight H. pylori-positive subjects between before and after H. pylori eradication (p=0.732, 0.618, and 0.435 in the antrum, body, and fundus, respectively), or between six eradicated and two noneradicated subjects (p=0.071, 0.857, 0.429, and 0.857 in the antrum, body, fundus, and plasma, respectively). CONCLUSIONS: These results show that H. pylori infection has no effect on stomach ghrelin levels and that eradication therapy does not influence plasma or tissue ghrelin levels.

A case of the primary malignant melanoma arising from uterine endometrium / 대한내과학회지

Malignant melanoma arising in the uterine endometrium is extremely rare. Only 12 cases of malignant melanoma of the uterine endometrium have been previously reported to date. All of them were metastatic cases. The most common presenting symptom was abnormal uterine bleeding. We report a case of primary malignant melanoma arising from the uterine endometrium. A 63-year-old multigravid woman presented with uterine bleeding. The pathologic review of an endometrial curettage specimen suggested an undifferentiated malignant tumor. A total abdominal hysterectomy, bilateral salpingo-oophorectomy, and pelvic lymph node dissection were performed. Final pathologic examination revealed malignant melanoma. Immunohistochemical studies demonstrated reactivity of tumor cells for HMB-45 and S-100 protein. She had no previous history of malignancy. Whole body PET scan was performed in an effort to find the primary site of malignant melanoma and showed no demonstrable abnormal FDG uptake suggesting malignancy elsewhere. This case is the first report of primary malignant melanoma involving uterine endometrium in Korea.

When should human papillomavirus (HPV) testing be done after conization? / 대한산부인과학회지

OBJECTIVE: To know when human papillomavirus (HPV) testing should be done after conization. METHODS: Between 1997 to 2004, Large Loop Excisions of the Transformation Zone (LLETZ) were done for conization to women with cervical pathology at A University Hospital. The Pap and HPV typing were done before LLETZ procedures. After conizations, HPV typing were planned to be done every 3 months. Every HPV typing was done by HPV oligonucleotide microarray (Biomedlab Co., Seoul, South Korea). RESULTS: For 8 years, 120 LLETZ were enrolled in this study. There were 8 cases of no neoplasm, 9 cases of CIN 1, 17 cases of CIN 2, 74 cases of CIN 3, 10 cases of microinvasive cervix cancer, and 2 cases of adenocarcinoma in situ. HPV DNA before LLETZ procedures was found about 85.0% and subtype 16 was the most common type among the patients with cervical lesion (40.8%). After LLETZ, 190 HPV typing were done through 1,307 total months (average, 6.9 months/typing). 95 (79.2%) cases had negative results, and 25 (20.8%) cases had positive results. Our data showed that, after conization, about 80% turned out to negative in 6 months. CONCLUSION: Our data suggested HPV DNA testing should be done after 6 months of LLETZ, as about 80% were destined to negative in 6 months.

Increased expression of pAKT is associated with radiation resistance in cervical cancer / 부인종양

OBJECTIVE: The aim of this study was to investigate the association of phosphorylated AKT (pAKT) expression and radiation resistance in cervical cancer. METHODS: A retrospective review was made of the records of 25 women who received primary radiation therapy due to locally advanced cervical cancer (LACC) with FIGO stage IIB-IVA. Nine patients regarded as radiation resistant developed local recurrences with a median progression free interval of 10 months. Sixteen patients did not show local recurrences, and were regarded as a radiation sensitive group. Using pretreatment paraffin-embedded tissues, we evaluated pAKT expression by immunohistochemistry. RESULTS: A significant association was found between the level of pAKT expression and local recurrence. Immunohistochemical staining for pAKT was significantly more frequent in the radiation resistant than in the radiation sensitive group (p=0.007). The mean progression free survival (PFS) was 84 months for patients with pAKT negative staining (17 cases) and 44 months for patients with pAKT positive expression (8 cases)(p=0.015). CONCLUSION: These results suggest that signaling from PI3K/pAKT can lead to radiation resistance in LACC.

High Frequency of Microsatellite Instability in Intestinal-type Gastric Cancer in Korean Patients

BACKGROUND: Although there have been some reports on microsatellite alterations in gastric cancer, findings are inconsistent regarding the associations between histological classification and microsatellite instability (MSI). In the present study, we attempted to determine whether Lauren's histological subtypes are related with MSI status. METHODS: Paraffin-embedded tissue samples from 14 diffuse-type and 14 intestinal-type gastric adenocarcinomas were matched up according to patient gender and age. Mononucleotide markers (BAT25 and BAT26) and dinucleotide markers (D2S123, D5S346, and D17S250) were used for MSI analyses. Microsatellite genotypes were categorized in terms of high MSI incidence (MSI-H, > 30% positive marker) or low MSI incidence (MSI-L, < 30% positive marker). Losses of hMLH1 and hMSH2 protein expression were immunohistochemically studied. RESULTS: MSI-H was observed in 11 cases (78%) of the 14 intestinal-type cases as compared to 3 (21%) of the 14 diffuse-type cases (p=0.007). In MSI-H tumors, 10 cases (71%) showed losses of hMLH1 protein expression, while 2 cases (14%) in MSI-L tumors showed losses of hMLH1 protein expression (p=0.006). CONCLUSION: MSI-H tumors are more frequently found in intestinal-type gastric cancer, which suggests the possibility that there are different pathogenic pathways in gastric carcinogenesis according to histologic type.

Decreased expression of galectin-3 is associated with the progression of cervical neoplasia / 부인종양

OBJECTIVE: Galectin-3, a member of the beta-galactoside-binding proteins, is an intracellular and extracellular lectin that interacts with intracellular glycoproteins, cell surface molecules and extracellular matrix proteins. Galectin-3 is expressed widely in epithelial and immune cells and the level of expression varies in many cancer cells relative to the normal tissues from which they arise. We investigated whether the expression of galectin-3 is associated with the progression of cervical neoplasia. METHODS: The galectin-3 expression was evaluated by immunohistochemistry in 90 formalin-fixed paraffin-embedded cervical tissues: 10 normal cervical specimens, 20 low-grade squamous intraepithelial lesions (LSILs), 20 high-grade squamous intraepithelial lesions (HSILs), and 40 invasive squamous cell carcinomas (ISCCs). RESULTS: The immunohistochemical staining showed that the expression of galectin-3 was strong in all normal cervical squamous epithelia. Staining gradually decreased in accordance with the histopathologic grades from an LSIL to an HSIL and an ISCC (P<0.001). In particular, the expression of galectin-3 was significantly decreased in HSILs (P=0.001) and this down-regulation was more pronounced in ISCCs than normal tissues (P<0.001). CONCLUSION: These data constitute the first observation that the expression of galectin-3 is down-regulated in cervical cancer tissues and suggest the decreased expression of this galactoside-binding lectin is associated with the progression of cervical neoplasia.

Expression of beta-catenin in Colorectal Cancer with Liver Metastasis

PURPOSE: Decreased expression of beta-catenin has been known to be associated with tumor metastasis. However, the clinical relationship between the degree of expression and the prognosis in colorectal cancer (CRC) remains unclear. In this study, we evaluated the prognostic value of beta-catenin expression in CRC patients with liver metastasis. METHODS: Paraffin embedded blocks were obtained from 70 patients who underwent potentially curative resection for CRC with liver metastasis. Samples from normal colon mucosa, primary CRC and metastatic liver lesion were prepared in tissue microarrays and were stained by immunohistochemistry with monoclonal antibody against beta- catenin. The membranous beta-catenin expression was assessed and the beta-catenin expression difference between primary CRC and metastatic liver lesion was analysed in relation to overall survival as well as disease free survival rates. RESULTS: In beta-catenin expression, preserved expression (score >6) was observed in 42.0%, and 21.9% of primary CRC tumor samples and tumor samples from metastatic liver lesion respectively. The degree of beta-catenin expression in metastatic liver lesion was significantly lower than that in primary CRC (P=0.022). According to the difference of beta-catenin expression score between primary CRC and liver metastasis, patients were classified as group 'A' and 'B'. Group 'A' was defined as patients showing remarkably decreased expression of beta-catenin in metastatic liver lesion in that the difference of the score was three or more. Group 'B' was defined as patients showing maintained or increased beta-catenin expression in metastatic liver lesion in comparison to primary CRC, in that the difference of beta-catenin expression score was less than three. Overall survival rate and disease free survival rate were significantly better in group 'B' than group 'A' (P=0.02, P=0.002). CONCLUSIONS: Decreased expression of beta-catenin in metastatic liver lesion may be a poor prognostic marker in colorectal cancers with liver metastasis. A further large-scaled investigation is necessary to define the role of beta-catenin in CRC.

Giant Multilocular Cystadenoma of the Prostate: A Case Report

Large multilocular cysts and cystic neoplasms of the prostate are very rare. A healthy 52-year-old man presented with acute urinary retension. Physical examination revealed a large abdominal mass. Pelvic MRI showed a 15x10x9 cm, lobulated, and multiseptated cystic mass in the pelvic cavity. Exploration laparotomy revealed a large cytic mass that compressd and displaced the urinary bladder antero-laterally, and adhered to the prostate and the sigmoid colon. Macroscopically, the mass was grayish white colored, multilocular cysts containing reddish-brown serous fluid. Microscopically, the tumor was composed of glands and cysts lined by the prostatic-type epithelium in the fibrous stroma. The prostatic nature of the lesion was confirmed by the lining epithelium expressing prostate-specific antigen.

Evaluation of Self-collected Pad Sampling for the Detection of HPV In Cervicovaginal Secretion

BACKGROUND: Self-collection of secretion samples for HPV testing is a feasible alternative method for women who would decline to participate in population based cervical cancer programs. The purpose of this study was to determine the sensitivity and specificity of self-sampling for HPV in determining high grade squamous intraepithelial lesion (HSIL) using the pad, and we also wished to compare the results from samples collected by women themselves and those results from samples collected by physicians. METHODS: Fifty patients voluntarily participated in the sensitivity and specificity study at the university hospitals and 290 volunteers participated in the agreement study at local clinics. DNA was extracted and amplified using HPV L1 consensus primers for the direct sequencing of the pad samples. RESULTS: For the detection of HSIL, self-collected pad sampling showed good sensitivity (75.0%) and excellent specificity (100%). Two hundreds eighty-six samples from the pads and concurrent physicians?samples showed the agreement at 98.6% with the Kappa, 0.9622 (p=0.0000). CONCLUSIONS: A self-sampling method using the pad for the detection of HPV DNA is suggested to be an efficient method to access many women for screening easily, rapidly and conveniently. Testing the pad method? utility for a country- or large area-based mass screening study will be necessary in the future.

Prognostic Value of VEGF in Human Pancreatic Ductal Adenocarcinoma

BACKGROUND: Since pancreatic cancer metastasizes early regardless of the size of the primary tumor, it is suggested that angiogenic factor is upregulated in this disease. Among the angiogenic factors, vascular endothelial growth factor (VEGF) is the most potent and specific growth factor. The aim of this study is to elucidate the prognostic value of VEGF expression in pancreatic cancers. METHODS: We analyzed the VEGF expression using immunohistochemistry in 72 resected pancreatic ductal adenocarcinomas. We examined the prognostic value of the VEGF expression along with its relationship with the clinicopathological features. RESULTS: VEGF expression and mutant p53 expression were not associated with microvessel density. VEGF expression was positively associated with mutant p53 expression. There were no statistically significant relationships between the VEGF expression and other clinicopathological features, such as age, sex, CA19-9, tumor size, location, tumor differentiation, and stage. VEGF expression was not associated with patient survival. CONCLUSION: VEGF expression was not associated with the microvessel density and patient survival in pancreatic ductal adenocarcinoma.

Prognostic Significance of Maspin in Pancreatic Ductal Adenocarcinoma

BACKGROUND: Maspin is a serpin family of protease inhibitors. Althouth maspin has been considered a tumor suppressor that inhibits the motility, invasion, and metastasis of breast and prostatic cancer cells, there are many conflicting reports about maspin expression and cancer prognosis. METHODS: To investigate whether the expression of maspin could be used as a prognostic marker in pancreatic cancer, 72 paraffin-embedded pancreatic ductal adenocarcinomas were analyzed using immunohistochemistry. We examined the prognostic value of maspin as well as its relationship with clinicopathological features. RESULTS: Maspin expression was observed in all pancreatic ductal adenocarcinoma. Unlike cancer tissues, however, faint or no expression was observed in the corresponding normal pancreatic tissues. In the Cox proportional hazard model, high maspin expression predicted a high hazard rate. Maspin expression had a positive correlation with tumor stage, but there were also no statistically significant relationships between maspin expression and other clinicopathological features. CONCLUSION: These findings suggest maspin expression to have biological relevance in the progression of pancreatic cancers, with potential use as a prognostic marker for pancreatic neoplasm with epithelial origin.