CAP TODAY Pathology/Laboratory Medicine/Laboratory Management
Editors: Michael Cibull, MD, professor emeritus, University of Kentucky College of Medicine, Lexington; Rouzan Karabakhtsian, MD, attending pathologist, Department of Pathology, Montefiore Medical Center, Albert Einstein College of Medicine, Bronx, NY; Thomas Cibull, MD, dermatopathologist, Evanston Hospital, NorthShore University HealthSystem, Evanston, Ill.; and Rachel Stewart, DO, resident physician, Department of Pathology and Laboratory Medicine, University of Kentucky.
Expanding the morphologic spectrum of differentiated VIN by mapping p53 loss
Mitotic count by PHH3 immunohistochemical staining in pancreatic WDNETs
Comparison of prostate cancer markers in lymph node and distant metastases
Evaluation of a new grading system for laryngeal squamous intraepithelial lesions
Breast cancer assessment based on levels of estrogen receptor expression
Validation of the VE1 immunostain for the BRAF V600E mutation in melanoma
Treatment-induced pathologic necrosis in STS after neoadjuvant chemoradiotherapy
Features of hepatocellular carcinoma arising in hepatocellular adenoma
Utility of re-excising mildly and moderately dysplastic nevi: a retrospective analysis
Expanding the morphologic spectrum of differentiated VIN by mapping p53 loss
The pathogenesis of vulvar squamous cell carcinoma follows one of two distinct pathways. A precursor lesion in the human papilloma virus-independent pathway—differentiated vulvar intraepithelial neoplasia (dVIN)—was only recently characterized in detail and is diagnosed infrequently without an associated component of invasive carcinoma. Aberrant p53 immunostaining is seen frequently in dVIN, and it manifests as a complete loss or a p53-null pattern in about 25 to 30 percent of cases. The abrupt transition between p53 loss and basal p53 expression in lesional versus nonlesional epithelium allows clear demarcation between neoplastic and non-neoplastic epithelium. The authors conducted a study focusing on the accuracy of diagnosis of dVIN. They identified for the study 14 specimens from 10 patients using the pathology archives of two teaching hospitals. The specimens were identified on the basis of a diagnosis of dVIN, with or without invasive carcinoma, and p53-null immunostaining pattern in lesional cells. Ten specimens had associated invasive carcinoma. All sections from each specimen that showed the specimen resection margin were stained for p53 and reviewed with all hematoxylin-and-eosin sections. Detailed morphologic assessment of the p53-null epithelium was made and compared with the adjacent benign squamous epithelium. The status of the resection margins based on the original pathologic assessment was compared with that assessed with p53 immunohistochemistry. One specimen showed p53 loss in the invasive carcinoma but patchy basal positivity in the region originally diagnosed as dVIN, supporting interpretation as a benign hyperplastic focus rather than dVIN. In the remaining 13 specimens, the areas originally diagnosed as dVIN, as well as the associated invasive carcinoma, if present, were p53-null. In eight of these specimens, dVIN was determined to be more extensive than originally recognized, based on the presence of p53-null immunostaining and subtle morphologic abnormalities. The spectrum of morphologic changes in p53-null regions that showed continuity with areas originally recognized as dVIN were subtle and typically consisted of an abrupt change in maturation of the squamous epithelium (loss of keratohyaline granules and parakeratosis); tinctorial alterations in the keratinocytes, with cells containing more abundant eosinophilic cytoplasm; and minimal basal nuclear atypia. Margin status changed from negative to positive in four of 13 specimens and from focally to more extensively positive in an additional three specimens. The authors concluded that better tools for the diagnosis of dVIN are needed. Until such tools are developed, the limitations in the current diagnosis of dVIN should be recognized.
Singh N, Leen SL, Han G, et al. Expanding the morphologic spectrum of differentiated VIN (dVIN) through detailed mapping of cases with p53 loss. Am J Surg Pathol. 2015;39:52–60.
Correspondence information not provided.
Mitotic count by PHH3 immunohistochemical staining in pancreatic WDNETs
Well-differentiated neuroendocrine tumors of the pancreas are graded on the basis of mitotic count or Ki67 index. Mitotic count has a narrow cutoff; its assessment is time consuming and carries poor interobserver reproducibility. Phosphohistone H3 (PHH3) is a mitosis-specific marker for which the value has been validated in several tumor types. The authors conducted a study to assess the utility of PHH3 in histologic grading of pancreatic well-differentiated neuroendocrine tumors (WDNETs). Sixty-three cases of surgically resected primary pancreatic WDNETs were retrieved, and immunohistochemical analysis for PHH3 and Ki67 was performed. Mitotic rate was independently assessed by four pathologists using hematoxylin and eosin (in 50 high-power fields [HPFs], expressed as mitoses per 10 HPFs) and PHH3 stains (in 50 HPFs, one 10×, and one 20× hotspot). PHH3 and Ki67 labeling indices were determined on a single 20× hotspot and expressed as the percentage of positive cells to total cells. Mitotic counts by various methods significantly correlated with each other and with PHH3 and Ki67 indices, with the best correlation seen within the three different PHH3 counts (in 50 HPFs, one 10×, and one 20× hotspot). Moreover, mitotic count on PHH3 was less time consuming than on hematoxylin and eosin (1.68 versus 3.67 minutes for 50 HPFs; P<0.0001). Histologic grade determined by PHH3 significantly correlated with disease-specific and disease-free survival, with the best cutoffs of ≥4 mitoses/10 HPFs (2 mm2), ≥7 mitoses/10× hotspot, ≥5 mitoses/20× hotspot (log rank test, P<0.0001), and ≥0.16 percent for PHH3 labeling index (log rank test, P<0.0006). Tumor grades based on PHH3 stain also showed significant correlation with patient survival in multivariate Cox proportional hazards models (P<0.05). Histologic grades by mitotic count on PHH3 demonstrated high concordance and kappa agreement with grades determined by mitotic count on hematoxylin and eosin. PHH3 stain also showed improved interobserver agreement in original mitotic count (intraclass correlation, 0.98 versus 0.79) and final grade assignment (Fleiss κ, 0.69 versus 0.46) as compared with hematoxylin and eosin. The authors concluded that their data confirmed that histologic grading by PHH3 stain has practical and prognostic value and offers reduced time and improved interobserver reproducibility in mitotic rate assessment and grade assignment. Although larger series are needed for validation, mitotic rate potentially can be determined by counting one hotspot, which will greatly facilitate the assessment of histologic grade in pancreatic WDNETs.
Voss SM, Riley MP, Lokhandwala PM, et al. Mitotic count by phosphohistone H3 immunohistochemical staining predicts survival and improves interobserver reproducibility in well-differentiated neuroendocrine tumors of the pancreas. Am J Surg Pathol. 2015;39:13–24.
Correspondence information not provided.
Comparison of prostate cancer markers in lymph node and distant metastases
Prostate cancer is primarily diagnosed at an early stage; however, some tumors are diagnosed in a metastatic stage as cancer of unknown primary origin. To allow specific treatment for prostate cancer presenting as cancer of unknown primary origin, it is important to determine the tumor’s origin. Prostate-specific antigen is used as a diagnostic marker for prostate cancer, but its expression declines with progression to castration-resistant prostate cancer. The authors conducted a study to identify the most informative marker constellation for detecting metastatic prostate cancer at high sensitivity. Widely used prostate cancer markers, such as prostate-specific antigen, prostate-specific acid phosphatase, androgen receptor, prostate-specific membrane antigen, prostein (also known as p501s or SLC45A3), and ETS-related gene, were investigated for their sensitivity in detecting the prostatic origin of metastases. Expression of prostate-specific antigen, prostate-specific acid phosphatase, androgen receptor, prostate-specific membrane antigen, prostein, and ETS-related gene was determined on archived tissue specimens consisting of benign prostatic tissue (n=9), primary prostate cancer (n=79), lymph node metastases (n=58), and distant metastases (n=39) using immunohistochemistry. Staining intensity was categorized as negative (zero), weak (one), moderate (two), and strong (three). All markers, except ETS-related gene, were able to detect at least 70 percent of lymph node metastases and distant metastases, with prostate-specific antigen, androgen receptor, and prostate-specific membrane antigen having the highest sensitivities (97 percent, 91 percent, and 94 percent, respectively). Sensitivity could be increased up to 98 percent and 100 percent by combining prostate-specific antigen, prostate-specific membrane antigen, or androgen receptor for lymph node metastases and distant metastases, respectively. The same level of sensitivity could be achieved by combining prostate-specific membrane antigen and prostein. These data show that combined staining of at least two prostate markers should be used to identify metastases as originating from prostate cancer.
Queisser A, Hagedorn SA, Braun M, et al. Comparison of different prostatic markers in lymph node and distant metastases of prostate cancer. Mod Pathol. 2015;28:138–145.
Correspondence: Dr. S. Perner at sven.perner1972@gmail.com
Evaluation of a new grading system for laryngeal squamous intraepithelial lesions
The authors conducted a study to verify the applicability, reproducibility, and predictive value of a proposed unified classification (amended Ljubljana classification) for laryngeal squamous intraepithelial lesions. Six internationally recognized experts and three pathologists from Ljubljana, Slovenia, contributed to the study by using the new system to evaluate a set of laryngeal squamous intraepithelial lesions, including low-grade SIL, high-grade SIL, and carcinoma in situ. The overall agreement among reviewers was good. Overall unweighted and weighted κ-values and 95 percent confidence intervals were 0.75 (0.65–0.84) and 0.80 (0.71–0.87), respectively. The results were stratified between the international reviewers and the Ljubljana pathologists. The former had good overall agreement, and the latter had very good agreement. Kaplan–Meier survival curves showed a significant difference (P<0.0001) between patients with low- and high-grade SILs: 19 of 1,204 patients with low-grade SILs and 30 of 240 patients with high-grade SILs progressed to malignancy in two to 15 years and two to 26 years, respectively. The authors concluded that the proposed modification to the Ljubljana classification provides clear morphological criteria for defining the prognostic groups. The criteria facilitate better interobserver agreement than previous systems, and the retrospective follow-up study demonstrated a highly significant difference in the risk of malignant progression between low- and high-grade SILs.
Gale N, Blagus R, El-Mofty SK, et al. Evaluation of a new grading system for laryngeal squamous intraepithelial lesions—a proposed unified classification. Histopathol. 2014;65:456–464.
Correspondence: Dr. N. Gale at nina.gale@mf.uni-lj.si
Breast cancer assessment based on levels of estrogen receptor expression
Historically, nuclear staining of 10 percent or more of invasive tumor cells has been used for estrogen receptor positivity. In 2010, a joint guideline from the American Society of Clinical Oncology and CAP recommended the cut-off value be changed to nuclear staining of one percent or more. The authors conducted a study to analyze the relationships between levels of estrogen receptor (ER) expression and clinicopathological features and clinical outcomes, with an emphasis on the ER expression one to 10 percent subgroup. They analyzed the clinicopathological features in five subgroups based on ER expression levels in 1,700 consecutive invasive breast cancer patients diagnosed and treated at their institution between 2000 and 2011. Of the cases, 24 percent had ER expression of less than one percent, two percent had ER of one to 10 percent, five percent had ER of 11 to 50 percent, five percent had ER of 51 to 70 percent, and 64 percent had ER of 71 to 100 percent. The authors observed four subgroups of patient cohorts (ER expression of less than one percent, one to 10 percent, 11 to 70 percent, and 71 to 100 percent) that were unique with regard to Nottingham grade, nuclear grade, progesterone receptor expression, and disease-free survival. For the 341 patients with follow-up data, they found no significant differences in pathological features between patients in the subgroups with ER expression of 11 to 50 percent and 51 to 70 percent. These data support the important role of ER in breast cancer, as well as the importance of accurate testing and quantitative reporting for ER. Tumors with ER expression of one to 10 percent are not common, and additional studies are needed to better understand this subgroup of breast cancer.
Zhang Z, Wang J, Skinner KA, et al. Pathological features and clinical outcomes of breast cancer according to levels of oestrogen receptor expression. Histopathol. 2014;65:508–516.
Correspondence: Dr. P. Tang at ping_tang@urmc.rochester.edu
Validation of the VE1 immunostain for the BRAF V600E mutation in melanoma
BRAF mutation status, and therefore eligibility for BRAF inhibitors, is determined by sequencing methods. The authors assessed the validity of VE1, a monoclonal antibody against the BRAF V600E mutant protein, in detecting mutant BRAF V600E melanomas as classified by DNA pyrosequencing. Their assessment included 76 metastatic melanoma patients with only one known primary melanoma who previously had BRAF codon 600 pyrosequencing of either their primary (n=19) or metastatic (n=57) melanoma, or both (n=17). All melanomas (n=93) were immunostained with the BRAF VE1 antibody using a red detection system. The staining intensity of these specimens was scored from 0 to 3+ by a dermatopathologist. Scores of 0 and 1+ were considered negative staining while scores of 2+ and 3+ were considered positive. The assessment found that VE1 antibody showed a sensitivity of 85 percent and specificity of 100 percent as compared to DNA pyrosequencing results. There was 100 percent concordance between VE1 immunostaining of primary and metastatic melanomas from the same patient. V600K, V600Q, and V600R BRAF melanomas did not stain positively with VE1. The authors concluded that this hospital-based study finds high sensitivity and specificity for the BRAF VE1 immunostain in comparison to pyrosequencing in detection of BRAF V600E in melanomas.
Pearlstein MV, Zedek DC, Ollila DW, et al. Validation of the VE1 immunostain for the BRAF V600E mutation in melanoma. J Cutan Pathol. 2014;41(9): 724–732.
Correspondence: Dr. Nancy Thomas at nthomas@med.unc.edu
Treatment-induced pathologic necrosis in STS after neoadjuvant chemoradiotherapy
Histologic response to chemotherapy has been shown to be an independent prognostic factor in patients with osteosarcoma and Ewing sarcoma. However, in patients with soft tissue sarcoma (STS), the prognostic impact of histologic response to chemotherapy is less clear. The authors sought to determine the prognostic significance of treatment-induced pathologic necrosis in patients receiving neoadjuvant chemoradiotherapy for STS. Between 1989 and 2011, they identified 113 patients with grade 2 or 3 (according to the National Cancer Institute grading system using three tiers) extremity or truncal STS who received neoadjuvant interdigitated chemoradiotherapy according to protocol followed by surgery. The extent of tumor necrosis in the resected specimens was quantified and correlated with outcome. The authors found that the median tumor necrosis rate was 90 percent, and 103 (91 percent) patients received all three cycles of planned neoadjuvant chemotherapy. The likelihood of achieving 95 percent or greater necrosis was not related to the number of preoperative cycles of chemotherapy received but was related to tumor histology (62 percent for malignant fibrous histiocytoma versus zero for synovial sarcoma [P<0.001]; 56 percent for myxoid liposarcoma versus zero for synovial sarcoma [P=0.002]). At a median follow-up of six years, no statistically significant differences were noted in the five-year local control, disease-specific survival, and overall survival rates for patients with 95 percent or greater necrosis (50 patients; 44 percent) and less than 95 percent necrosis (63 patients; 56 percent), even when stratifying by histology. The authors concluded that in a homogeneous population of patients with high-grade extremity and truncal STS who were treated with neoadjuvant chemoradiotherapy, the extent of pathologic tumor necrosis did not correlate with outcome.
Mullen JT, Hornicek FJ, Harmon DC, et al. Prognostic significance of treatment-induced pathologic necrosis in extremity and truncal soft tissue sarcoma after neoadjuvant chemoradiotherapy. Cancer. 2014;120:3676–3682.
Correspondence: Dr. John T. Mullen at jmullen@partners.org
Features of hepatocellular carcinoma arising in hepatocellular adenoma
Well-differentiated hepatocellular carcinoma in noncirrhotic liver can show morphological features similar to hepatocellular adenoma. In rare instances, hepatocellular carcinoma can arise in the setting of hepatocellular adenoma. The authors conducted a study to compare the immunohistochemical and cytogenetic features of the hepatocellular adenoma-like and hepatocellular carcinoma portions of these tumors. Immunohistochemistry for β-catenin, glutamine synthetase, serum amyloid A protein, glypican-3, and heat-shock protein 70 was performed in 11 cases of hepatocellular carcinoma arising in hepatocellular adenoma in noncirrhotic liver. Tumors with nuclear β-catenin or diffuse glutamine synthetase, or both, were considered β-catenin activated. Fluorescence in situ hybridization (FISH) was performed in nine cases for gains of chromosomes 1, 8, and MYC. The study involved seven men (33 to 75 years old) and four women (29 to 65 years old). Focal atypical morphological features were seen in hepatocellular adenoma-like areas in seven (64 percent) cases. Hepatocellular adenoma-like areas showed features of inflammatory hepatocellular adenoma in seven (64 percent) cases; and four were also serum amyloid A positive in the hepatocellular carcinoma portion. β-catenin activation, heat-shock protein 70 positivity, and chromosomal gains on FISH were seen in the hepatocellular adenoma portion in 55 percent, 40 percent, and 56 percent of cases, and 73 percent, 60 percent, and 78 percent of cases in the hepatocellular carcinoma portion, respectively. The authors concluded that the hepatocellular adenoma-like portion of most cases of hepatocellular carcinoma arising in hepatocellular adenoma shows features typically seen in hepatocellular carcinoma, such as focal morphological abnormalities, β-catenin activation, heat-shock protein 70 expression, and chromosomal gains. Hepatocellular adenoma-like areas in these tumors, especially in men and older women, may represent an extremely well-differentiated variant of hepatocellular carcinoma, whereas the morphologically recognizable hepatocellular carcinoma portion represents a relatively higher grade component of the tumor.
Kakar S, Grenert JP, Paradis V, et al. Hepatocellular carcinoma arising in adenoma: similar immunohistochemical and cytogenetic features in adenoma and hepatocellular carcinoma portions of the tumor. Mod Pathol. 2014;27:1499–1509.
Correspondence: Dr. S. Kakar at sanjay.kakar@ucsf.edu
Utility of re-excising mildly and moderately dysplastic nevi: a retrospective analysis
The management of dysplastic nevi is a highly debated and controversial topic within the dermatology community. Clinicians agree that margin-positive severely dysplastic nevi (DN) should be removed with a surgical margin; however, there is disagreement surrounding the appropriate management of this type of mole. The authors evaluated the utility of re-excising margin-positive mildly and moderately DN. They conducted a retrospective chart review on all adult patients given the diagnosis of a biopsy-proven DN from 2010 through 2011. The primary outcomes were defined as the presence of melanocytic residuum in re-excisional specimens and a clinically significant change in diagnosis. A total of 1,809 mildly and moderately DN were diagnosed from 2010 through 2011. In all, 765 (42.3 percent) of these lesions were found to have positive surgical margins during biopsy, and 495 (64.7 percent) of the 765 lesions were subsequently re-excised. Melanocytic residuum was present in 18.2 percent of re-excisional specimens. Re-excision resulted in a clinically significant alteration of the diagnosis in only one case (0.2 percent). The authors acknowledged that their study was limited by its retrospective design and an inability to assess for malignant transformation given limited follow-up. They concluded that re-excising mildly and moderately DN results in a low histopathological yield and rarely results in a clinically significant change in diagnosis. Consequently, clinical monitoring of margin-positive lesions may be warranted.
Strazzula L, Vedak P, Hoang MP, et al. The utility of re-excising mildly and moderately dysplastic nevi: a retrospective analysis. J Am Acad Dermatol. 2014;71:1071–1076.
Correspondence: Dr. Daniela Kroshinsky, Dept. of Dermatology, Massachusetts General Hospital, 50 Staniford St., #200, Boston, MA 02114