IU Indianapolis ScholarWorks :: Browsing by Subject "immunohistochemistry"

Browsing by Subject "immunohistochemistry"

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Analytical validation of a standardised scoring protocol for Ki67 immunohistochemistry on breast cancer excision whole sections: an international multicentre collaboration
(Wiley, 2019-08) Leung, Samuel C. Y.; Nielsen, Torsten O.; Zabaglo, Lila A.; Arun, Indu; Badve, Sunil S.; Bane, Anita L.; Bartlett, John M. S.; Borgquist, Signe; Chang, Martin C.; Dodson, Andrew; Ehinger, Anna; Fineberg, Susan; Focke, Cornelia M.; Gao, Dongxia; Gown, Allen M.; Gutierrez, Carolina; Hugh, Judith C.; Kos, Zuzana; Lænkholm, Anne-Vibeke; Mastropasqua, Mauro G.; Moriya, Takuya; Nofech-Mozes, Sharon; Osborne, C. Kent; Penault-Llorca, Frédérique M.; Piper, Tammy; Sakatani, Takashi; Salgado, Roberto; Starczynski, Jane; Sugie, Tomoharu; van der Vegt, Bert; Viale, Giuseppe; Hayes, Daniel F.; McShane, Lisa M.; Dowsett, Mitch; Pathology and Laboratory Medicine, School of Medicine
Aims The nuclear proliferation marker Ki67 assayed by immunohistochemistry has multiple potential uses in breast cancer, but an unacceptable level of interlaboratory variability has hampered its clinical utility. The International Ki67 in Breast Cancer Working Group has undertaken a systematic programme to determine whether Ki67 measurement can be analytically validated and standardised among laboratories. This study addresses whether acceptable scoring reproducibility can be achieved on excision whole sections. Methods and results Adjacent sections from 30 primary ER+ breast cancers were centrally stained for Ki67 and sections were circulated among 23 pathologists in 12 countries. All pathologists scored Ki67 by two methods: (i) global: four fields of 100 tumour cells each were selected to reflect observed heterogeneity in nuclear staining; (ii) hot‐spot: the field with highest apparent Ki67 index was selected and up to 500 cells scored. The intraclass correlation coefficient (ICC) for the global method [confidence interval (CI) = 0.87; 95% CI = 0.799–0.93] marginally met the prespecified success criterion (lower 95% CI ≥ 0.8), while the ICC for the hot‐spot method (0.83; 95% CI = 0.74–0.90) did not. Visually, interobserver concordance in location of selected hot‐spots varies between cases. The median times for scoring were 9 and 6 min for global and hot‐spot methods, respectively. Conclusions The global scoring method demonstrates adequate reproducibility to warrant next steps towards evaluation for technical and clinical validity in appropriate cohorts of cases. The time taken for scoring by either method is practical using counting software we are making publicly available. Establishment of external quality assessment schemes is likely to improve the reproducibility between laboratories further.
Analytical validation of a standardized scoring protocol for Ki67: phase 3 of an international multicenter collaboration
(Nature, 2016) Leung, Samuel C. Y.; Nielsen, Torsten O.; Zabaglo, Lila; Arun, Indu; Badve, Sunil S.; Bane, Anita L.; Bartlett, John M. S.; Borgquist, Signe; Chang, Martin C.; Dodson, Andrew; Enos, Rebecca A.; Fineberg, Susan; Focke, Cornelia M.; Gao, Dongxia; Gown, Allen M.; Grabau, Dorthe; Gutierrez, Carolina; Hugh, Judith C.; Kos, Zuzana; Lænkholm, Anne-Vibeke; Lin, Ming-Gang; Mastropasqua, Mauro G.; Moriya, Takuya; Nofech-Mozes, Sharon; Osborne, C. Kent; Penault-Llorca, Frédérique M.; Piper, Tammy; Sakatani, Takashi; Salgado, Roberto; Starczynski, Jane; Viale, Giuseppe; Hayes, Daniel F.; McShane, Lisa M.; Dowsett, Mitch; Pathology and Laboratory Medicine, School of Medicine
Pathological analysis of the nuclear proliferation biomarker Ki67 has multiple potential roles in breast and other cancers. However, clinical utility of the immunohistochemical (IHC) assay for Ki67 immunohistochemistry has been hampered by unacceptable between-laboratory analytical variability. The International Ki67 Working Group has conducted a series of studies aiming to decrease this variability and improve the evaluation of Ki67. This study tries to assess whether acceptable performance can be achieved on prestained core-cut biopsies using a standardized scoring method. Sections from 30 primary ER+ breast cancer core biopsies were centrally stained for Ki67 and circulated among 22 laboratories in 11 countries. Each laboratory scored Ki67 using three methods: (1) global (4 fields of 100 cells each); (2) weighted global (same as global but weighted by estimated percentages of total area); and (3) hot-spot (single field of 500 cells). The intraclass correlation coefficient (ICC), a measure of interlaboratory agreement, for the unweighted global method (0.87; 95% credible interval (CI): 0.81–0.93) met the prespecified success criterion for scoring reproducibility, whereas that for the weighted global (0.87; 95% CI: 0.7999–0.93) and hot-spot methods (0.84; 95% CI: 0.77–0.92) marginally failed to do so. The unweighted global assessment of Ki67 IHC analysis on core biopsies met the prespecified criterion of success for scoring reproducibility. A few cases still showed large scoring discrepancies. Establishment of external quality assessment schemes is likely to improve the agreement between laboratories further. Additional evaluations are needed to assess staining variability and clinical validity in appropriate cohorts of samples.
Antiarrhythmic effects of stimulating the left dorsal branch of the thoracic nerve in a canine model of paroxysmal atrial tachyarrhythmias
(Elsevier, 2018) Zhao, Ye; Yuan, Yuan; Tsai, Wei-Chung; Jiang, Zhaolei; Tian, Zhi-peng; Shen, Changyu; Lin, Shien-Fong; Fishbein, Michael C.; Everett, Thomas H., IV.; Chen, Zhenhui; Chen, Peng-Sheng; Medicine, School of Medicine
Background Stellate ganglion nerve activity (SGNA) precedes paroxysmal atrial tachyarrhythmia (PAT) episodes in dogs with intermittent high-rate left atrial (LA) pacing. The left dorsal branch of the thoracic nerve (LDTN) contains sympathetic nerves originating from the stellate ganglia. Objective The purpose of this study was to test the hypothesis that high-frequency electrical stimulation of the LDTN can cause stellate ganglia damage and suppress PAT. Methods We performed chronic LDTN stimulation in 6 dogs with and 2 dogs without intermittent rapid LA pacing while monitoring SGNA. Results LDTN stimulation reduced average SGNA from 4.36 μV (95% confidence interval [CI] 4.10–4.62 μV) at baseline to 3.22 μV (95% CI 3.04–3.40 μV) after 2 weeks (P = .028) and completely suppressed all PAT episodes in all dogs studied. Tyrosine hydroxylase staining showed large damaged regions in both stellate ganglia, with increased percentages of tyrosine hydroxylase–negative cells. The terminal deoxynucleotidyl transferase dUTP nick end labeling assay showed that 23.36% (95% CI 18.74%–27.98%) of ganglion cells in the left stellate ganglia and 11.15% (95% CI 9.34%–12.96%) ganglion cells in the right stellate ganglia were positive, indicating extensive cell death. A reduction of both SGNA and heart rate was also observed in dogs with LDTN stimulation but without high-rate LA pacing. Histological studies in the latter 2 dogs confirmed the presence of extensive stellate ganglia damage, along with a high percentage of terminal deoxynucleotidyl transferase dUTP nick end labeling–positive cells. Conclusion LDTN stimulation damages both left stellate ganglia and right stellate ganglia, reduces left SGNA, and is antiarrhythmic in this canine model of PAT.
Distinct clinicopathological features in metanephric adenoma harboring BRAF mutation
(Impact Journals, 2016-07-08) Caliò, Anna; Eble, John N.; Hes, Ondrej; Martignoni, Guido; Harari, Saul E.; Williamson, Sean R.; Brunelli, Matteo; Osunkoya, Adeboye O.; Wang, Lisha; Comperat, Eva; Lopez-Beltran, Antonio; Wang, Mingsheng; Zhang, Shaobo; Curless, Kendra L.; Post, Kristin M.; Chang, Hsim-Yee; Luchini, Claudio; Baldrige, Lee Ann; MacLennan, Gregory T.; Montironi, Rodolfo; Grignon, David J.; Cheng, Liang; Pathology and Laboratory Medicine, School of Medicine
BRAF mutation recently has been reported in metanephric adenoma. We sought to determine the clinical and morphologic features of BRAF-mutated metanephric adenoma and to correlate BRAF mutation with BRAF V600E immunohistochemical staining results. A series of 48 metanephric adenomas and 15 epithelial-predominant nephroblastomas were analyzed for the occurrence of BRAF mutation (BRAF V600E/V600E complex, BRAF V600D, BRAF V600K and BRAF V600R) using the BRAF RGQ PCR kit (Qiagen). Immunohistochemistry was performed using monoclonal mouse antibodies against p16INK4 and VE1 (Spring Bioscience), recognizing the BRAF V600E mutant protein. Forty-one of 48 cases (85%) showed BRAF V600E mutation; none of the other BRAF variants was detected. Of 41 BRAF-mutated metanephric adenomas, 33 showed positive VE1 immunostaining (sensitivity 80%, specificity 100%); in all cases we detected p16INK4 expression regardless of BRAF mutation status. All epithelial-predominant nephroblastomas were BRAF-wild-type and none expressed VE1. The following features were associated with BRAF V600E mutation: older patients (p=0.01), female predominance (p=0.005) and the presence of a predominantly acinar architecture (p=0.003). In summary, BRAF-mutated metanephric adenomas were associated with older age, female predominance, and the presence of a predominant acinar component. A subset (20%) of BRAF-mutated metanephric adenomas was not detected by VE1 immunostaining.
ESR1 and PGR polymorphisms are associated with estrogen and progesterone receptor expression in breast tumors
(American Physiological Society, 2016-09-01) Hertz, Daniel L.; Henry, N. Lynn; Kidwell, Kelley M.; Thomas, Dafydd; Goddard, Audrey; Azzouz, Faouzi; Speth, Kelly; Li, Lang; Banerjee, Mousumi; Thibert, Jacklyn N.; Kleer, Celina G.; Stearns, Vered; Hayes, Daniel F.; Skaar, Todd C.; Rae, James M.; Medicine, School of Medicine
Hormone receptor-positive (HR+) breast cancers express the estrogen (ERα) and/or progesterone (PgR) receptors. Inherited single nucleotide polymorphisms (SNPs) in ESR1, the gene encoding ERα, have been reported to predict tamoxifen effectiveness. We hypothesized that these associations could be attributed to altered tumor gene/protein expression of ESR1/ERα and that SNPs in the PGR gene predict tumor PGR/PgR expression. Formalin-fixed paraffin-embedded breast cancer tumor specimens were analyzed for ESR1 and PGR gene transcript expression by the reverse transcription polymerase chain reaction based Oncotype DX assay and for ERα and PgR protein expression by immunohistochemistry (IHC) and an automated quantitative immunofluorescence assay (AQUA). Germline genotypes for SNPs in ESR1 (n = 41) and PGR (n = 8) were determined by allele-specific TaqMan assays. One SNP in ESR1 (rs9322336) was significantly associated with ESR1 gene transcript expression (P = 0.006) but not ERα protein expression (P > 0.05). A PGR SNP (rs518162) was associated with decreased PGR gene transcript expression (P = 0.003) and PgR protein expression measured by IHC (P = 0.016), but not AQUA (P = 0.054). There were modest, but statistically significant correlations between gene and protein expression for ESR1/ERα and PGR/PgR and for protein expression measured by IHC and AQUA (Pearson correlation = 0.32–0.64, all P < 0.001). Inherited ESR1 and PGR genotypes may affect tumor ESR1/ERα and PGR/PgR expression, respectively, which are moderately correlated. This work supports further research into germline predictors of tumor characteristics and treatment effectiveness, which may someday inform selection of hormonal treatments for patients with HR+ breast cancer.
Identification of novel biomarker candidates for immunohistochemical diagnosis to distinguish low-grade chondrosarcoma from enchondroma
(Wiley, 2015-07) Lai, Xianyin; Chen, Shaoxiong; Department of Biochemistry and Molecular Biology, IU School of Medicine
Chondrosarcoma is the third most common primary bone cancer, requiring surgical resection. However, differentiation of low-grade chondrosarcoma (grade 1) from enchondroma that is benign and only requires regular follow-up is one of the most frequent diagnostic dilemmas facing orthopedic oncologists in clinical management. Although multiple techniques are applied to make the distinction, immunohistochemistry is an important ancillary technique, especially when a histopathological stain of specimen must be obtained in order to guarantee an accurate confirmation. Currently, no adequate immunohistochemical diagnostic protein biomarkers are available to distinguish low-grade chondrosarcoma from enchondroma. To discover novel protein biomarker candidates, an LC-MS/MS approach was applied to directly compare formalin-fixed, paraffin-embedded low-grade chondrosarcoma with enchondroma tissue samples. The proteomics analysis revealed 17 protein biomarker candidates. A principle was developed to prioritize the candidates using category and ranking. An algorithm, prioritization index of biomarker candidates for immunohistochemistry on tissue specimens, was developed to rank the candidates inside each category. Using the proteomics data and bioinformatics results, the prioritization index of biomarker candidates for immunohistochemistry on tissue revealed periostin as a top candidate. Immunohistochemical staining of periostin in 23 low-grade chondrosarcoma and 31 enchondroma tissue specimens disclosed 87% specificity and 70% sensitivity.
Immunophenotypic and pathologic heterogeneity of unclassified renal cell carcinoma: a study of 300 cases
(Elsevier, 2020-08) Akgul, Mahmut; Cheng, Liang; Urology, School of Medicine
Renal cell carcinoma, unclassified (RCC-U), is a heterogenous group of tumors that do not fit in any of the recognized histologic types. Immunohistochemical studies are frequently used to characterize these tumors. Herein, we sought to investigate the immunophenotypes of 300 cases of RCC-U. The cases were morphologically classified into three groups: oncocytoma/chromophobe renal cell carcinoma–like, group 1; clear cell renal cell carcinoma–like, group 2; and others (ie, papillary renal cell carcinoma–like/collecting duct–like/pure sarcomatoid), group 3. The male-to-female ratio was 1.4. Most cases (168, 66%) were group 1. Group 3 was associated with larger tumor size, advanced stage, and frequent lymph node metastases. The most commonly used immunohistochemical stains were CK7 (n = 270; 89.5%), vimentin (n = 186, 82%), CD10 (n = 181; 59.9%), and AMACR (n = 162; 54%). Pancytokeratin (79/101; 78.2%) and PAX8 (54/61; 88.5%) were diffusely positive in most cases, followed by AMACR (69/117; 59%). CD117 was positive in 53 of 118 cases (45%). RCC-U is a morphologically and immunophenotypically heterogenous group of tumors, and comprehensive workup is needed before rendering the diagnosis.
Immunophenotypic Comparison of Testicular Sclerosing Sertoli Cell Tumors and Sertoli Cell Tumors Not Otherwise Specified
(Elsevier, 2017-09) Mesa, Hector; Zhang, Chen; Manivel, Juan C.; Ulbright, Thomas M.; Department of Pathology and Laboratory Medicine, School of Medicine
Testicular Sertoli cell tumors (SCTs) are rare, and most fall into the category of SCT–not otherwise specified (SCT-NOS). Only a few additional types of SCT are recognized. Sclerosing SCT (S-SCT), originally described in 1991, comprises a small fraction of SCTs and was considered a specific entity until the 2016 revision of the World Health Organization classification of non–germ cell tumors, where it was classified as a morphologic variant of SCT-NOS. In a recent study, differences in expression of PAX2/PAX8, inhibin, androgen receptor, and S100 protein between SCT-NOS and S-SCT were noted in a small number of cases. In this interinstitutional study, we compared the expression of these markers and β-catenin in 11 cases each of SCT-NOS and S-SCT to determine if differences exist that could justify keeping a separate classification of these neoplasms. PAX2/PAX8 cocktail was the only marker that was significantly overexpressed in S-SCT. Expression of androgen receptors was strong in S-SCT and variable in SCT-NOS but did not reach statistical significance. Expression of β-catenin was common in both, whereas inhibin was infrequent. The available material was insufficient for a conclusive evaluation of S100 protein expression. Overall, our results support the inclusion of S-SCT as a morphologic variant of SCT-NOS. Expression of PAX2/PAX8 in S-SCT may reflect an overactive epithelial-to-mesenchymal transition as has been shown in experimental models of acute and chronic seminiferous tubular injury and might be related to the process generating the stroma in these tumors.
Prognostic role of Ape/Ref-1 subcellular expression in stage I-III breast carcinomas
(2002-01) Puglisi, Fabio; Barbone, Fabio; Tell, Gianluca; Aprile, Giuseppe; Pertoldi, Barbara; Raiti, Concetta; Kelley, Mark R.; Damante, Giuseppe; Sobrero, Alberto; Beltrami, Carlo Alberto; Di Loreto, Carla
The purpose of the study was to evaluate the prognostic role of the DNA repair protein APE/Ref-1 in breast carcinomas. Immunohistochemical analysis for APE/Ref-1 was performed in a series of 133 consecutive stage I-III breast carcinomas. The relationship between APE/Ref-1 and other prognostic and predictive factors such as tumor size, nodal status, histologic grade, p53 expression, hormonal receptor status, vascular invasion and necrosis was investigated. The prognostic value of APE/Ref-1 was studied by univariate and multivariate analysis. The predominant pattern of APE/Ref-1 immunohistochemical expression was nuclear, although cytoplasmic and mixed nuclear/cytoplasmic localization was also observed. The percentage of cells with APE/Ref-1 cytoplasmic stain directly correlated with the percentage of p53 positive cases (rho=0.28, p=0.013). The small group of women whose tumors showed mixed nuclear/cytoplasmic APE/Ref-1 localization (n=5) experienced a significantly poorer survival (p=0.014) and Cox proportional hazard model analysis identified APE/Ref-1 as an independent prognostic factor. The results suggest that subcellular localisation of APE/Ref-1 may influence the aggressiveness of breast carcinomas.
Report From the International Society of Urological Pathology (ISUP) Consultation Conference on Molecular Pathology of Urogenital Cancers: IV: Current and Future Utilization of Molecular-Genetic Tests for Testicular Germ Cell Tumors
(Wolters Kluwer, 2020-03-20) Looijenga, Leendert H. J.; Van der Kwast, Theodorus H.; Grignon, David; Egevad, Lars; Kristiansen, Glen; Kao, Chia-Sui; Idrees, Muhammad T.; Pathology and Laboratory Medicine, School of Medicine
The International Society of Urological Pathology (ISUP) organized a Consultation Conference in March 2019 dealing with applications of molecular pathology in Urogenital Pathology, including testicular tumors (with a focus on germ cell tumors [GCTs]), preceded by a survey among its members to get insight into current practices in testicular germ cell tumor (TGCT) diagnostics and adoption of the ISUP immunohistochemical guidelines published in 2014. On the basis of the premeeting survey, the most commonly used immunomarker panel includes OCT3/4, placental alkaline phosphate, D2-40, SALL4, CD117, and CD30 for GCTs and the documentation of germ cell neoplasia in situ (GCNIS). Molecular testing, specifically 12p copy gain, is informative to distinguish non-GCNIS versus GCNIS related GCTs, and establishing germ cell origin of tumors both in the context of primary and metastatic lesions. Other molecular methodologies currently available but not widely utilized for TGCTs include genome-wide and targeted approaches for specific genetic anomalies, P53 mutations, genomic MDM2 amplification, and detection of the p53 inactivating miR-371a-3p. The latter also holds promise as a serum marker for malignant TGCTs. This manuscript provides an update on the classification of TGCTs, and describes the current and future role of molecular-genetic testing. The following recommendations are made: (1) Presence of GCNIS should be documented in all cases along with extent of spermatogenesis; (2) Immunohistochemical staining is optional in the following scenarios: identification of GCNIS, distinguishing embryonal carcinoma from seminoma, confirming presence of yolk sac tumor and/or choriocarcinoma, and differentiating spermatocytic tumor from potential mimics; (3) Detection of gain of the short arm of chromosome 12 is diagnostic to differentiate between non-GCNIS versus GCNIS related GCTs and supportive to the germ cell origin of both primary and metastatic tumors.

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