SP142 PD-L1 Scoring Shows High Interobserver and Intraobserver Agreement in Triple-negative Breast Carcinoma But Overall Low Percentage Agreement With Other PD-L1 Clones SP263 and 22C3.

SP142 programmed cell death ligand 1 (PD-L1) status predicts response to atezolizumab in triple-negative breast carcinoma (TNBC). Prevalence of VENTANA PD-L1 (SP142) Assay positivity, concordance with the VENTANA PD-L1 (SP263) Assay and Dako PD-L1 IHC 22C3 pharmDx assay, and association with clinicopathologic features were assessed in 447 TNBCs. SP142 PD-L1 intraobserver and interobserver agreement was investigated in a subset of 60 TNBCs, with scores enriched around the 1% cutoff. The effect of a 1-hour training video on pretraining and posttraining scores was ascertained. At a 1% cutoff, 34.2% of tumors were SP142 PD-L1 positive. SP142 PD-L1 positivity was significantly associated with tumor-infiltrating lymphocytes (P <0.01), and node negativity (P=0.02), but not with tumor grade (P=0.35), tumor size (P=0.58), or BRCA mutation (P=0.53). Overall percentage agreement (OPA) for intraobserver and interobserver agreement was 95.0% and 93.7%, respectively, among 5 pathologists trained in TNBC SP142 PD-L1 scoring. In 5 TNBC SP142 PD-L1-naive pathologists, significantly higher OPA to the reference score was achieved after video training (posttraining OPA 85.7%, pretraining OPA 81.5%, P<0.05). PD-L1 status at a 1% cutoff was assessed by SP142 and SP263 in 420 cases, and by SP142 and 22C3 in 423 cases, with OPA of 88.1% and 85.8%, respectively. The VENTANA PD-L1 (SP142) Assay is reproducible for classifying TNBC PD-L1 status by trained observers; however, it is not analytically equivalent to the VENTANA PD-L1 (SP263) Assay and Dako PD-L1 IHC 22C3 pharmDx assay.

The International Academy of Cytology Yokohama System for Reporting Breast Fine Needle Aspiration Biopsy Cytopathology: Analysis and discussion of the response to a web-based survey.

BACKGROUND: A group of international experts in breast fine needle aspiration biopsy (FNAB) cytopathology, supported by the International Academy of Cytology (IAC), drafted a comprehensive system for reporting breast FNAB cytopathology in 2017-2018. The editorial team produced a survey to assess the international response to the proposed category structure, definitions, and management recommendations in this draft. METHODS: A web-based survey of 186 questions was generated using the Qualtrics software package (Provo, Utah) supported by the Division of Information Technology at the University of Wisconsin-Madison. The survey was advertised widely-including through the IAC, American Society of Cytopathology, Japanese Society of Clinical Cytology, Papanicolaou Society of Cytopathology, and Australian Society of Cytology and to audiences at national and international meetings-and was available from April to June 2018. The data obtained from the 265 respondents was assessed by the editorial team. RESULTS: The survey provided a snapshot of the current role and use of breast FNAB and the international variations. Demographic questions were followed by specific questions based on the draft category definitions and statements and focused on issues that had generated discussion among the authors, including the FNAB diagnosis of ductal carcinoma in situ. CONCLUSION: The survey results strongly supported the development of the IAC Yokohama System and informed subsequent discussions among the authors regarding the final text.

Discordance in 21-gene recurrence scores between paired breast cancer samples is inversely associated with patient age.

BACKGROUND: The Oncotype DX 21-gene Recurrence Score is a genomic-based algorithm that guides adjuvant chemotherapy treatment decisions for women with early-stage, oestrogen receptor (ER)-positive breast cancer. However, there are age-related differences in chemotherapy benefit for women with intermediate Oncotype DX Recurrence Scores that are not well understood. Menstrual cycling in younger women is associated with hormonal fluctuations that might affect the expression of genomic predictive biomarkers and alter Recurrence Scores. Here, we use paired human breast cancer samples to demonstrate that the clinically employed Oncotype DX algorithm is critically affected by patient age. METHODS: RNA was extracted from 25 pairs of formalin-fixed paraffin-embedded, invasive ER-positive breast cancer samples that had been collected approximately 2 weeks apart. A 21-gene signature analogous to the Oncotype DX platform was assessed through quantitative real-time PCR, and experimental recurrence scores were calculated using the Oncotype DX algorithm. RESULTS: There was a significant inverse association between patient age and discordance in the recurrence score. For every 1-year decrease in age, discordance in recurrence scores between paired samples increased by 0.08 units (95% CI - 0.14, - 0.01; p = 0.017). Discordance in recurrence scores for women under the age of 50 was driven primarily by proliferation- and HER2-associated genes. CONCLUSION: The Oncotype DX 21-gene Recurrence Score algorithm is critically affected by patient age. These findings emphasise the need for the consideration of patient age, particularly for women younger than 50, in the development and application of genomic-based algorithms for breast cancer care.

Breast fine needle aspiration biopsy cytology: the potential impact of the International Academy of Cytology Yokohama System for Reporting Breast Fine Needle Aspiration Biopsy Cytopathology and the use of rapid on-site evaluation.

The present report reviews the current problems associated with the routine use of breast fine needle aspiration biopsy (FNAB) and discusses the potential impact that the new International Academy of Cytology (IAC) Yokohama Reporting System and the use of rapid on-site evaluation (ROSE) should have on reducing these problems to optimize breast care for patients. The recently reported IAC System aims to establish the best practice guidelines for breast FNAB, emphasizing the importance of the FNAB technique and the skillful preparation of direct smears. The IAC System proposes a standardized report and established clear terminology for defined reporting categories, each of which has a risk of malignancy and is linked to management options. The FNAB techniques that will optimize the biopsy specimen and reduce poor quality smears are reviewed and the benefits of ROSE are discussed. FNAB can diagnose accurately the vast majority of breast lesions, and ROSE has been recommended whenever possible to reduce the rate of insufficient/inadequate cases and increase the number of specific benign and malignant diagnoses. ROSE performed by a cytopathologist provides a provisional diagnosis, reducing patient anxiety and facilitating management through cost-effective immediate triage and patient selection for ancillary testing. Thus, patients can be selected for immediate core needle biopsy, as required.

The International Academy of Cytology Yokohama System for Reporting Breast Fine-Needle Aspiration Biopsy Cytopathology.

The International Academy of Cytology (IAC) gathered together a group of cytopathologists expert in breast cytology who, working with clinicians expert in breast diagnostics and management, have developed the IAC Yokohama System for Reporting Breast Fine-Needle Aspiration Biopsy (FNAB) Cytology. The project was initiated with the first cytopathology group meeting in Yokohama at the 2016 International Congress of Cytology. This IAC Yokohama System defines five categories for reporting breast cytology, each with a clear descriptive term for the category, a definition, a risk of malignancy (ROM) and a suggested management algorithm. The key diagnostic cytopathology features of each of the lesions within each category will be presented more fully in a subsequent atlas. The System emphasizes that the crucial requirements for diagnostic breast FNAB cytology are a high standard for the performance of the FNAB and for the making of direct smears, and well-trained experienced cytopathologists to interpret the material. The performance indicators of breast FNAB, including specificity and sensitivity, negative predictive value, positive predictive value and ROM stated in this article have been derived from the recent literature. The current practice of breast FNAB has evolved with the increasing use of ultrasound guidance and rapid on-site evaluation. Two recent publications have shown a range of ROM for the insufficient/inadequate category of 2.6-4.8%, benign 1.4-2.3%, atypical 13-15.7%, suspicious of malignancy 84.6-97.1%, and malignant 99.0-100%. The management algorithm in the System provides options because there are variations in the management of breast lesions using FNAB and core-needle biopsy in those countries utilizing the "triple test" of clinical, imaging, and FNAB assessment, and also variations in the availability of CNB and imaging in low- and middle-income countries. The System will stimulate further discussion and research, particularly in the cytological diagnostic features of specific lesions within each category and in management recommendations. This will lead to continuing improvements in the care of patients with breast lesions and possible modifications to the IAC Yokohama System.

Ductal carcinoma in situ: breast-conserving surgery without radiotherapy.

BACKGROUND: Ductal carcinoma in situ (DCIS) is commonly managed with breast-conserving surgery (BCS) and adjuvant radiotherapy. Oncoplastic BCS allows wide excision without compromising the breast's natural shape. We use 'level one' techniques to excise a 'sector' of tissue (apex at the nipple) rather than traditional 'lumpectomy'. There are concerns that some DCIS is over-treated and radiotherapy administered unnecessarily incurring the associated cost, time and morbidity without added benefit. This study aims to determine if pure DCIS can be managed safely without relying upon adjuvant therapies with an acceptable breast conservation rate. METHOD: A retrospective clinical study of 96 patients who underwent BCS alone for pure DCIS between 1995 and 2009. Data were collected on patient's demographics, presentation, excision margins, re-excision and final margins, size, grade and nuclear architecture of DCIS, recurrent ipsilateral breast events, contralateral breast events, date of the last follow-up with mammography and patients' assessment of cosmetic outcome (scale 1-5: very dissatisfied, somewhat dissatisfied, somewhat satisfied, satisfied, very satisfied). RESULTS: Twelve (12.5%) had an ipsilateral recurrence. Mean follow-up was 7.6 years (median 7.4); yearly recurrence rate was 1.67%. Of the 52 patients who gave an assessment of cosmetic outcome, 46 were satisfied. CONCLUSION: Ipsilateral recurrence rates were favourable compared with previous trials where DCIS was treated with lumpectomy and radiotherapy. Oncoplastic techniques may be used to successfully treat pure DCIS with BCS alone without adjuvant therapies.

Assessing HER2 amplification in breast cancer: findings from the Australian In Situ Hybridization Program.

In August 2006, the Australian government approved subsidized trastuzumab therapy for human epidermal growth factor receptor 2 (HER2)-positive early breast cancer, and it was mandated that HER2 testing should be performed using in situ hybridization (ISH) rather than immunohistochemistry (IHC). Here we review results of the first regulated, nationwide program to provide HER2 ISH testing for all newly diagnosed breast cancer patients, with a particular emphasis on cases where IHC and ISH results were discordant. Data from all laboratories participating in the program were collated. Cases with an equivocal ISH test result [by chromogenic ISH (CISH) or silver ISH (SISH)] were tested centrally by fluorescence ISH. Most laboratories also performed HER2 IHC, and 200 cases with discordant IHC and ISH results were selected for further analysis in a central laboratory. A total of 26 laboratories were involved and 53,402 tests were reported. Over a 4-year period the HER2 positivity rate decreased for primary cancers from 23.8 to 14.6 %, but remained relatively constant for samples from metastases. Average ISH reporting times were <5 days for all yearly reporting periods. Test-repeat rates decreased for CISH (8.9-3.6 %) and SISH (13.7-8.4 %). Only 12 of 196 cases remained discordant after retesting in a central laboratory. These findings demonstrate the successful implementation of a regulated, national program that continues to collect data on HER2 status. The results also highlight the differences in IHC interpretation between local laboratories and a central, more experienced, laboratory. This model could be used to establish future biomarker-testing programs in other countries.

Gastric HER2 Testing Study (GaTHER): an evaluation of gastric/gastroesophageal junction cancer testing accuracy in Australia.

Trastuzumab provides a survival benefit in patients with human epidermal growth factor receptor 2 (HER2)-amplified/overexpressed advanced gastric and gastroesophageal junction cancers (GC/GJCs). However, the optimal method for testing is unclear. The aim of this study was to assess interlaboratory agreement on HER2 scoring in GC/GJC to aid the development of a robust testing algorithm for diagnostic practice in Australia. Nine laboratories assessed the HER2 status of 100 GC/GJC tissue samples by immunohistochemistry (IHC) and in situ hybridization (ISH) [chromogenic (CISH) or silver (SISH)] using both HER2 copy number and HER2:chr17 (chromosome 17) ratio. Results were compared with reference fluorescence ISH (FISH). Interlaboratory agreement on IHC3+ scoring was good (κ=0.76), and there was good/very good agreement between IHC (positivity defined as IHC3+) and ISH when HER2 copy number was used (κ=0.72 to 0.87). Agreement on CISH/SISH scoring was good/very good when HER2 copy number was used (κ=0.68 to 0.86), and agreement between CISH/SISH and FISH using HER2 copy number was very good (κ=0.88 to 0.91). Agreement was reduced when HER2:chr17 ratio was used. The good agreement for HER2 copy number determined by bright-field ISH suggests that this is the optimal method for testing in GC/GJC cases. An IHC3+ score was strongly predictive of a positive ISH result, although agreement for all IHC scores was only moderate, suggesting that IHC triage before ISH testing would be the most cost-effective strategy. However, because of the unique features of GC/GJC samples and the difficulty of ensuring consistent HER2 staining in the community setting, it is recommended that HER2 status in advanced GC/GJC be determined by both IHC and ISH in the same laboratory.

Establishment of the Australian in situ hybridization program for the assessment of HER2 amplification in breast cancer: a model for the introduction of new biomarkers into clinical practice.

In August 2006, the Australian government announced a decision to subsidize trastuzumab therapy for early breast cancer, to commence 6 weeks later. It was mandated that HER2 gene amplification, determined by in situ hybridization (ISH), be shown, and that the sponsor company, Roche Products Pty Ltd, should fund this testing. This announcement potentially required provision of ISH testing for HER2 for every newly diagnosed breast cancer, where previously HER2 testing had been performed by immunohistochemistry with support from a single fluorescence ISH (FISH) reference laboratory for indeterminate cases. The Australian HER2 Testing Advisory Board, an independent expert group, responded to the challenge of rapidly providing accurate nationwide ISH testing. Bright-field ISH was selected as the testing platform and a decentralized testing model, with support from a central FISH laboratory, was adopted. An implementation plan was developed addressing standards for training, accreditation, and quality assurance. Within 6 weeks, 8 pathology laboratories were accredited for ISH testing and by September 2008, 2 years after the announcement, 22 ISH testing laboratories were taking part in the national program and almost 20,000 ISH tests had been performed. This article describes the design and rapid implementation of a nationwide program of bright-field ISH as the first-line testing platform for HER2 status in early breast cancer. We believe that this model for the coordinated and large-scale implementation of a new biomarker test has wide application, given that accurate assessment of a range of novel biomarkers is being used increasingly to determine eligibility for new targeted treatment modalities.

Global levels of specific histone modifications and an epigenetic gene signature predict prostate cancer progression and development.

BACKGROUND: Epigenetic alterations are common in prostate cancer, yet how these modifications contribute to carcinogenesis is poorly understood. We investigated whether specific histone modifications are prognostic for prostate cancer relapse, and whether the expression of epigenetic genes is altered in prostate tumorigenesis. METHODS: Global levels of histone H3 lysine-18 acetylation (H3K18Ac) and histone H3 lysine-4 dimethylation (H3K4diMe) were assessed immunohistochemically in a prostate cancer cohort of 279 cases. Epigenetic gene expression was investigated in silico by analysis of microarray data from 23 primary prostate cancers (8 with biochemical recurrence and 15 without) and 7 metastatic lesions. RESULTS: H3K18Ac and H3K4diMe are independent predictors of relapse-free survival, with high global levels associated with a 1.71-fold (P < 0.0001) and 1.80-fold (P = 0.006) increased risk of tumor recurrence, respectively. High levels of both histone modifications were associated with a 3-fold increased risk of relapse (P < 0.0001). Epigenetic gene expression profiling identified a candidate gene signature (DNMT3A, MBD4, MLL2, MLL3, NSD1, and SRCAP), which significantly discriminated nonmalignant from prostate tumor tissue (P = 0.0063) in an independent cohort. CONCLUSIONS: This study has established the importance of histone modifications in predicting prostate cancer relapse and has identified an epigenetic gene signature associated with prostate tumorigenesis. IMPACT: Our findings suggest that targeting the epigenetic enzymes specifically involved in a particular solid tumor may be a more effective approach. Moreover, testing for aberrant expression of epigenetic genes such as those identified in this study may be beneficial in predicting individual patient response to epigenetic therapies.

Bilateral atypical ductal hyperplasia, an incidental finding in gynaecomastia--case report and literature review.

Male breast cancer is a rare disease. Atypical ductal hyperplasia (ADH) in men is much rarer, and bilateral involvement is exceptional. A 20-year-old male presented with bilateral gynaecomastia who underwent subcutaneous mastectomies and histopathology revealed bilateral ADH. At 24 months, completion mastectomies were performed on both sides. The residual breast tissue revealed ADH similar to the initial specimen. ADH in women increases the risk of breast cancer by four to five times. To our knowledge, this is the first case report of bilateral ADH in a gynaecomastia specimen.

Expression of extracellular matrix components versican, chondroitin sulfate, tenascin, and hyaluronan, and their association with disease outcome in node-negative breast cancer.

PURPOSE: The purpose is to determine whether the levels of expression of extracellular matrix components in peritumoral stroma are predictive of disease outcome for women with node-negative breast cancer. EXPERIMENTAL DESIGN: Tumor tissue from 86 patients with node-negative breast cancer was examined by immunohistochemical staining for the expression of versican, chondroitin sulfate (CS), tenascin, and hyaluronan (HA). With the exception of HA, the expression of the extracellular matrix components was measured by video image analysis. Statistical correlation of the immunohistochemical data with clinicopathological characteristics and disease outcome was performed. RESULTS: All of the extracellular matrix components were present in the peritumoral stroma of the entire study cohort. In contrast, immunoreactivity within the cancer cell was observed in 82% of tumors for HA, 12% for CS, and 4% for tenascin; no immunostaining of cancer cells for versican was observed for any of the tumors. Cox regression and Kaplan-Meier analyses indicated that elevated expression of stromal versican predicted increased risk and rate of relapse in this cohort. Elevated expression of tenascin was predictive of increased risk and rate of death only. Although neither CS nor HA were predictive of disease outcome in this cohort, tumor size was predictive of increased risk and rate of both relapse and survival. CONCLUSIONS: Elevated expression within peritumoral stromal matrix of versican and tenascin was predictive of relapse-free and overall survival, respectively, in women with node-negative breast cancer.

Expression and localization of homeodomain proteins DLX4/HB9 in normal and malignant human breast tissues.

BACKGROUND: Alterations in the control of gene expression is a key event in neoplastic transformation. Investigating the expression of transcription factors such as homeodomain proteins may therefore allow better characterization of molecular mechanisms underlying the transformation process. MATERIALS AND METHODS: Expression of homeodomain proteins DLX4 and HB9 was detected by RT-PCR and immunohistochemically in 24 breast tumors and their corresponding non-malignant tissue. RESULTS: Although the percentage of nuclei expressing both DLX4 (p = 0.001) and HB9 (p = 0.0001) is increased in breast carcinoma, their intensity of nuclear staining is decreased compared to non-malignant nuclei. Furthermore HB9 nuclear immunoreactivity decreased progressively with increasing tumor grade (p = 0.001). CONCLUSION: These data suggest that the reduction in malignant nuclear DLX4 immunoreactivity is an earlier event in breast carcinogenesis than the progressive loss of HB9 expression observed with increasing tumor grade. With further study and in conjunction with standard criterion of pathology, these findings may help to predict the malignant behaviour of some breast cancers.

Regulation of stromal versican expression by breast cancer cells and importance to relapse-free survival in patients with node-negative primary breast cancer.

PURPOSE: Determination of meaningful prognostic indicesremains a high priority for women diagnosed with node-negative primary breast cancer. Currently, 30% of these women relapse, and there is no reliable means of predicting this group of patients. This study investigates whether the level of expression of versican, an anticell adhesive proteoglycan, in the peritumoral stromal tissue of women with node-negative, primary breast cancer predicts relapse-free survival. This study also examines whether breast cancer cells regulate the secretion of versican by mammary fibroblasts. EXPERIMENTAL DESIGN: Immunoreactive versican was measured in breast cancer tissue sections of 58 node-negative patients by video image analysis. Primary isolates of mammary fibroblasts were cultured in medium conditioned by the breast cancer cell lines ZR-75-1, MCF-7, BT-20, and MB231. Changes in versican secretion were measured by immunoblotting and enhanced chemiluminescence. RESULTS: Cox analyses indicated that peritumoral versican level was the sole predictor of relapse-free survival. The relapse rate in patients with low versican levels was lower than in patients with high versican levels (Kaplan-Meier: 83% relapse free at 5 years for versican mean integrated absorbance <14 versus 33% for > or = 14, P = 0.0006). Accumulation of versican in medium of mammary fibroblasts was increased after culture in conditioned medium from breast cancer cell lines. CONCLUSIONS: Relapse in women with node-negative breast cancer is related to the level of versican deposited in peritumoral stroma by mammary fibroblasts. Versican secretion appears to be regulated by breast cancer cell mediators. Neoplastic remodeling of extracellular matrix through increased versican deposition may facilitate local invasion and metastasis.