Detailed DNA methylation characterisation of phyllodes tumours identifies a signature of malignancy and distinguishes phyllodes from metaplastic breast carcinoma.

Phyllodes tumours (PTs) are rare fibroepithelial lesions of the breast that are classified as benign, borderline, or malignant. As little is known about the molecular underpinnings of PTs, current diagnosis relies on histological examination. However, accurate classification is often difficult, particularly for distinguishing borderline from malignant PTs. Furthermore, PTs can be misdiagnosed as other tumour types with shared histological features, such as fibroadenoma and metaplastic breast cancers. As DNA methylation is a recognised hallmark of many cancers, we hypothesised that DNA methylation could provide novel biomarkers for diagnosis and tumour stratification in PTs, whilst also allowing insight into the molecular aetiology of this otherwise understudied tumour. We generated whole-genome methylation data using the Illumina EPIC microarray in a novel PT cohort (n = 33) and curated methylation microarray data from published datasets including PTs and other potentially histopathologically similar tumours (total n = 817 samples). Analyses revealed that PTs have a unique methylome compared to normal breast tissue and to potentially histopathologically similar tumours (metaplastic breast cancer, fibroadenoma and sarcomas), with PT-specific methylation changes enriched in gene sets involved in KRAS signalling and epithelial-mesenchymal transition. Next, we identified 53 differentially methylated regions (DMRs) (false discovery rate < 0.05) that specifically delineated malignant from non-malignant PTs. The top DMR in both discovery and validation cohorts was hypermethylation at the HSD17B8 CpG island promoter. Matched PT single-cell expression data showed that HSD17B8 had minimal expression in fibroblast (putative tumour) cells. Finally, we created a methylation classifier to distinguish PTs from metaplastic breast cancer samples, where we revealed a likely misdiagnosis for two TCGA metaplastic breast cancer samples. In conclusion, DNA methylation alterations are associated with PT histopathology and hold the potential to improve our understanding of PT molecular aetiology, diagnostics, and risk stratification. © 2024 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Hibernoma: a case report of a rare cardiac tumour.

Background: A cardiac hibernoma is a rare phenomenon, with just a handful of reports in the literature. They are difficult to characterize with conventional imaging including echocardiography, computed tomography (CT), cardiac magnetic resonance (CMR), or positron emission tomography (PET). Their definitive diagnosis relies primarily on histopathology via either endovascular or surgical biopsy. Previous case reports have entailed surgical excision followed by histopathology; however, surgery may be unfavourable in some patients with increased perioperative risk. Case summary: We present the case of a 57-year-old woman who was referred to our cardiology service with an interatrial lipomatous mass found incidentally on chest CT for assessment of rib fractures. She had 6 months of unexplained syncope, which was attributed to superior vena cava (SVC) compression demonstrated by chest CT. The mass had benign characteristics on echocardiography, CT, and CMR but was glucose-avid on PET, which indicated a possible malignancy such as liposarcoma. Her comorbid and very significant airways disease precluded her from surgical excision, so instead, endovascular biopsy was performed. Histopathology showed brown fat which was negative for mouse double minute 2 amplification on fluorescence in situ hybridisation testing; hence, a diagnosis was made of hibernoma, a rare benign tumour of brown fat. Given the benign diagnosis and her surgical risk with severe chronic obstructive pulmonary disease, a multidisciplinary recommendation was made favouring conservative management, with careful ongoing follow-up and the consideration of SVC stenting if symptoms progressed. Discussion: The definitive diagnosis of a cardiac hibernoma is complex and relies heavily on histopathology due to the contradictory findings on chest imaging. Careful consideration of management within a multidisciplinary team setting is essential to achieve a successful outcome.

Looking beyond workforce parity: addressing gender inequity in pathology.

While women pathologists have made up over one-third of pathologists in the Australian workforce for over 15 years and at least 50% since 2019, they are under-represented in senior leadership roles, scientific publications, grant recipients, editorial boards, key presentations, and professional awards. This is not unique to pathology and is seen in the broader medical and academic community. Barriers to gender equity and equality in pathology, medicine and academia include gender stereotypes, gender-based discrimination, structural and organisational barriers as well as broader social and cultural barriers. A diverse leadership reflective of the whole professional body and the broader community is important for optimal health outcomes. It is the responsibility and moral duty of individuals and organisations to address any gender disparities, inequities, and inequalities by monitoring, identifying, and acting on gender biases and systemic barriers that hinder appropriate levels of representation by women.

Inter-rater concordance of basal cell carcinoma subtypes: influences on reporting format and opportunities for further classification modifications.

Diagnosis of basal cell carcinoma (BCC) higher risk subtypes influences management strategies because of their propensity to recur locally. Subtyping is prone to inter-observer variability, and subtyping definitions are inconsistently applied. This study sought to compare the interobserver reproducibility of individual BCC subtypes using the 4th edition World Health Organization (WHO) Classification of Skin Tumours (CoST) definitions, with classification into lower and higher risk histological subtype groups. Ninety-one BCC cases were rated by seven pathologists, noting the presence of BCC subtype(s), and providing a higher or lower risk subtype grouping per case. Raters were provided with definitions as per the 4th edition WHO CoST for 10 listed BCC subtypes. Surgical specimen type was noted. Subgroup analysis was performed to exclude cases when the tumour deep front was not well visualised, or there was tangential sectioning (n = 6). Light's kappa was used to assess inter-rater reliability. From the total group (n = 91), five BCC subtypes showed a sufficient number of ratings for computing a κ statistic. From these five subtypes, superficial subtype showed substantial inter-rater agreement (κ = 0.64), and the other four subtypes showed moderate inter-rater agreement [nodular (κ = 0.45), sclerosing/morphoeic (κ = 0.45), infiltrating (κ = 0.49) and micronodular (κ = 0.57)]. Two-tiered rating into either higher or lower risk subtype showed substantial inter-rater agreement (κ = 0.72). Our results suggest a need to more precisely define BCC subtypes. We suggest reporting BCC subtype using a two-tiered risk grouping, followed by specific subtypes present. Further studies examining the inter-rater reliability of less common BCC subtypes are required.

Mesenchymal chondrosarcoma: An Australian multi-centre cohort study.

BACKGROUND: Mesenchymal chondrosarcoma (MCS) is an ultra-rare sarcoma that follows a more aggressive course than conventional chondrosarcoma. This study evaluates prognostic factors, treatments (surgery, chemotherapy, and radiation), and outcomes in an Australian setting. METHODS: We collected demographics, clinicopathological variables, treatment characteristics, and survival status from patients with MCS registered on the national ACCORD sarcoma database. Outcomes include overall survival (OS) and progression-free survival (PFS). RESULTS: We identified 22 patients with MCS between 2001-2022. Median age was 28 (range 10-59) years, 19 (86%) had localised disease at diagnosis of whom 16 had surgery (84%), 11 received radiation (58%), and 10 chemotherapy (53%). Ten (52%) developed recurrence and/or metastases on follow-up and three patients with initial metastatic disease received surgery, radiation, and chemotherapy. At a median follow-up of 50.9  (range 0.4-210) months nine patients had died. The median OS was 104.1 months (95% CI 25.8-182.3). There was improved OS for patients with localised disease who had surgical resection of the primary (p = 0.003) and those with ECOG 0-1 compared to 2-3 (p = 0.023) on univariate analysis. CONCLUSIONS: This study demonstrates contemporary Australian treatment patterns of MCS. The role of chemotherapy for localised disease remains uncertain. Understanding treatment patterns and outcomes help support treatment decisions and design of trials for novel therapeutic strategies.

NKX3.1 immunohistochemistry is highly specific for the diagnosis of mesenchymal chondrosarcomas: experience in the Australian population.

Mesenchymal chondrosarcoma (MC) is a rare sarcoma that typically arises in adolescents and young adults and characteristically harbours a HEY1-NCOA2 gene fusion. A recent study has shown that NKX3.1 immunohistochemistry (IHC) is highly specific and sensitive in MCs. NKX3.1 is a nuclear marker expressed in prostatic tissue and is widely used in most laboratories to determine prostatic origin of metastatic tumours. In the current study we investigated whether this stain can be used in the diagnostic workup of MC, as it may assist in triaging cases for further molecular testing, by assessing its expression in a cohort of MCs and in a wide spectrum of sarcoma types. Furthermore, we aimed to elucidate if expression of NKX3.1 by MCs is related to androgen receptor (AR) expression. We identified NKX3.1 positive nuclear staining in 9 of 12 individual patients of MC (n=20 of 25 samples when taking into account separate episodes). Four of the five negative specimens had been previously subjected to acid-based decalcification. NKX3.1 was negative in 536 samples from 16 non-MC sarcomas derived from largely tissue microarrays (TMAs). Overall, we identified 80% sensitivity and 100% specificity for NKX3.1 IHC in MCs. The sensitivity increased to 95.2% when acid-based decalcified specimens were excluded from the analysis. No correlation between NKX3.1 expression and AR IHC was identified. In summary, our findings indicate that NKX3.1 nuclear positivity is highly sensitive and specific for MC, provided that ethylenediaminetetraacetic acid (EDTA)-based rather than acid-based decalcification is used for sample processing. NKX3.1 IHC in the right clinical and histopathological setting can potentially be sufficient for the diagnosis of MC, reserving molecular confirmation only for equivocal cases.

PD-1 blockade using pembrolizumab in adolescent and young adult patients with advanced bone and soft tissue sarcoma.

BACKGROUND: Sarcomas represent 10%-15% of cancers in adolescent and young adult (AYA) patients, and survival for those with metastatic disease or relapse is poor. Immunotherapy with checkpoint inhibition has improved outcomes in multiple tumor types, but data in advanced sarcomas, particularly within the AYA population, are limited. AIM: We aim to evaluate response and toxicity for AYA patients with sarcoma treated with pembrolizumab. METHODS AND RESULTS: We retrospectively reviewed AYA patients with advanced bone and soft tissue sarcoma who received self-funded pembrolizumab between May 2015 and January 2019. Eighteen patients were identified. One patient with Ewing sarcoma had a sustained complete response to therapy. Two patients with alveolar soft part sarcoma received a clinical benefit from pembrolizumab: one had a radiological partial response with an excellent clinical response and one patient achieved stable disease. Four patients died of disease prior to first scheduled assessment and thus were not evaluable. The remaining eleven patients had progressive disease. CONCLUSION: The role of immunotherapy in AYA sarcoma warrants further investigation. Biomarkers of response need to be further evaluated in order to guide patient selection.

Primary dermal melanoma: clinical behaviour, prognosis and treatment.

PURPOSE: Primary dermal melanoma (PDM) is a subtype of cutaneous melanoma, confined to the dermis, which poses a challenging clinical dilemma. It may represent a true primary melanoma or a dermal cutaneous metastasis. This study aimed to delineate the histopathological characteristics and prognosis of PDM in a large patient cohort to guide appropriate treatment strategies. METHODS: A search of the Melanoma Research Database at Melanoma Institute Australia was conducted to identify all possible PDM patients at our institution diagnosed from 1978 to 2013. Overall, melanoma-specific and disease-free survival outcomes of the PDM group were compared to those of similar cohorts of Stage I-II and Stage IV M1a melanoma patients based on propensity score matching. RESULTS: Sixty-two PDM patients were identified from the MRD with a median follow-up of 6.3 years. Five-year survival was 87.1% and overall survival was 74.2%. PDMs had a significantly improved overall survival (p = 0.0002) and melanoma-specific survival (p = 0.001) compared to Stage I-II controls, however there was no difference in disease-free survival (p = 0.08). PDMs also demonstrated improved overall survival (p < 0.0001), melanoma-specific survival (p < 0.0001) and disease-free survival (p < 0.0001) compared to Stage IV M1a controls. CONCLUSION: These findings demonstrate that PDMs have a more favorable prognosis compared to stage I-II cutaneous melanomas and suggest that these are in fact true primary lesions. This study thus provides evidence to justify a treatment approach, by way of a wide local excision and possibly sentinel lymph node biopsy, as for early stage primary cutaneous melanomas.

The prognostic significance of microsatellites in cutaneous melanoma.

Microscopic satellite metastases are an adverse prognostic feature in primary cutaneous melanoma patients. The prognostic significance of microsatellites, including their number, size and distance from the primary melanoma, using the 8th edition American Joint Committee on Cancer definition, has not previously been evaluated. This study sought to determine the prognostic significance of microsatellites in histopathologically reviewed cases. Eighty-seven cases of primary cutaneous melanoma with the presence of microsatellites documented in the original pathology report and all histopathology slides available were reviewed and the findings were correlated with clinical outcome. Matched control cases were selected for all confirmed microsatellites cases. The presence of microsatellites was confirmed in 69 cases. The microsatellite group had significantly worse prognosis, with 21% 5-year disease-free survival compared with 56% in the control group (p < 0.001). The 5-year melanoma-specific survival was 53% in the microsatellites group and 73% in the control group (p = 0.004). Increasing distance (mm) of the microsatellite from the primary melanoma was found to adversely influence disease-free survival (HR = 1.24, 95% CI: 1.13-1.36, p < 0.001), overall survival (HR = 1.26 95%CI: 1.13-1.40, p < 0.001), and melanoma-specific survival (HR = 1.27 95% CI: 1.11-1.45, p < 0.001). Number and size of microsatellites were not significant prognostic factors. The presence of microsatellites was the only factor that proved to be an independent predictor of sentinel node positivity in multivariate analysis (OR 4.64; 95% CI 1.66-12.95; p = 0.003). Microsatellites were significantly associated with more loco-regional recurrences (p < 0.001) but not distant metastases (p = 0.821). Melanomas with microsatellites as defined by the 8th edition American Joint Committee on Cancer staging system are thus aggressive tumors, associated with significantly worse disease-free survival, overall survival and melanoma-specific survival. The presence of microsatellites is also associated with sentinel node-positivity and local and in-transit recurrence. Increasing distance of the microsatellite from the primary tumor is an independent adverse prognostic factor that warrants further evaluation.

FISH analysis of selected soft tissue tumors: Diagnostic experience in a tertiary center.

AIM: Fluorescence in situ hybridization (FISH) is an important ancillary tool for the classification of bone/soft tissue (BST) tumors. The aim of this study was to evaluate the contribution of FISH to the final classification of common BST entities in the molecular pathology department of the Royal Prince Alfred Hospital (RPAH), which is one of the most important referral centers for the management of sarcomas in Australia. METHODS: All routine diagnostic FISH tests performed on BST formalin-fixed paraffin embedded (FFPE) tissue specimens at the RPAH in a 5-year period (February, 2010-November, 2015) were reviewed. FISH analyses presented in this study include commercial break-apart probes (SS18, FUS, DDIT3, FUS, USP6, PDGFB, TFE3 and ALK) and a single enumeration (MDM2) probe. RESULTS: There were 434 interpretable FISH assays on BST samples including MDM2 (n=180), SS18 (n=97), FUS (n=64), DDIT3 (n=37), USP6 (n=30), PDGFB (n=13), TFE3 (n=8) and ALK (n=5). Discrepancies between the histopathological diagnosis and the FISH results were seen in 12% of the cases. In this subset of discordant cases, FISH contributed to the re-classification of 7% of cases originally diagnosed as synovial sarcoma (SS18) and 6% of adipocytic neoplasms (MDM2) based on the presence or absence of the expected gene alteration. CONCLUSION: Our study confirms that paraffin FISH is a sensitive and specific ancillary tool in the diagnosis of BST neoplasms when used in the appropriate clinicopathological context. These findings highlight the need for further ancillary molecular tools in the diagnosis and characterization of challenging cases.

Cutaneous presentation of gastrointestinal adenocarcinoma.

Seeding of a central nervous system malignancy to the abdominal cavity is an uncommon but well documented complication of a ventriculoperitoneal (VP) shunt. However, the metastasis of a primary gastrointestinal cancer to the skin via a VP shunt is extremely rare. We report the clinical case of an 85-year-old male who presented with a right upper quadrant nodule over his shunt, which on histopathology and tumour marker profile was diagnosed as an adenocarcinoma of likely upper gastrointestinal origin. This case illustrates the importance of proceeding to biopsy to inform prognosis and management, despite the risks of shunt infection.

Atypical Ewing sarcoma breakpoint region 1 fluorescence in-situ hybridization signal patterns in bone and soft tissue tumours: diagnostic experience with 135 cases.

AIMS: Recurrent Ewing sarcoma breakpoint region 1 (EWSR1) gene rearrangements characterize a select group of bone and soft tissue tumours. In our routine diagnostic practice with fluorescence in-situ hybridization (FISH), we have occasionally observed EWSR1 gene rearrangements in tumours not associated classically with EWSR1 translocations. This study aimed to review our institutional experience of this phenomenon and also to highlight the occurrence of unusual EWSR1 FISH signals (i.e. 5' centromeric region or 3' telomeric region signals) that do not fulfil the published diagnostic criteria for rearrangements. METHODS AND RESULTS: Using an EWSR1 break-apart probe, we performed FISH assays on formalin-fixed paraffin-embedded tissue sections from 135 bone and soft tissue specimens as part of their routine diagnostic work-up. EWSR1 gene rearrangements were identified in 51% of cases, 56% of which also showed an abnormal FISH signal pattern (in addition to classically rearranged signals). However, atypical FISH signals were present in 45% of the non-rearranged cases. In addition, we observed tumours unrelated to those described classically as EWSR1-associated that were technically EWSR1-rearranged in 6% of cases. Borderline levels of rearrangement (affecting 10-30% of lesional cells) were present in an additional 17% of these cases. CONCLUSIONS: While our study confirmed that FISH is a sensitive and specific tool in the diagnosis of EWSR1-associated tumours, atypical FISH signals and classical rearrangement in entities other than EWSR1-associated tumours can occur. Therefore, it is essential that the FISH result not be used as an isolated test, but must be evaluated in the context of clinical features, imaging, pathological and immunohistochemical findings.

Pathology review significantly affects diagnosis and treatment of melanoma patients: an analysis of 5011 patients treated at a melanoma treatment center.

BACKGROUND: Pathologists sometimes disagree on the diagnosis of melanoma or its histopathologic staging, which may have implications for treatment and follow-up. For this reason, melanoma patients referred to Melanoma Institute Australia (MIA) for further treatment routinely have their pathology slides reviewed by MIA pathologists. This study sought to determine whether diagnosis, staging, and treatment of melanoma patients changed significantly after central pathology review. METHODS: A total of 5,011 pairs of non-MIA and MIA pathology reports on the same primary melanoma specimen were reviewed. Differences in diagnosis, American Joint Committee on Cancer (AJCC) T classification, and treatment recommendations based on the non-MIA and MIA pathology reports were determined. RESULTS: A melanoma diagnosis changed in 5.1 % of cases after review. Where both pathologists agreed on a diagnosis of melanoma, AJCC T classification changed in 22.1 % after review. After MIA review, planned surgical excision margins changed in 11.2 % of cases, and a recommendation for sentinel lymph node biopsy (SLNB) changed in 8.6 %. Non-MIA reports less frequently contained criteria to define AJCC T classification (86.6 vs. 97.6 %), select appropriate surgical excision margins (95.2 vs. 99.6 %) and make a recommendation for SLNB (94.5 vs. 99.4 %), (each p < 0.001). On multivariate analysis, partial biopsies were independently associated with more frequent changes in AJCC T classification (p < 0.001), planned surgical excision margins (p < 0.001), and SLNB recommendations (p < 0.001) on the basis of MIA pathology review. CONCLUSIONS: Diagnosis, AJCC T classification, and treatment recommendations often change after pathology review by specialist melanoma pathologists. We recommend pathology review be considered for all patients attending specialist melanoma treatment centers.

Reproducibility of AJCC staging parameters in primary cutaneous melanoma: an analysis of 4,924 cases.

BACKGROUND: Pathology reports are of critical importance for conveying information to clinicians who must make important management decisions for their patients. This study sought to assess and compare the precision, reproducibility, and completeness of external pathology reports and pathology reports generated by central review of each case in a large cohort of primary cutaneous melanoma patients. METHODS: Details of matched external pathology reports and corresponding review reports for 4,924 primary cutaneous invasive melanomas diagnosed and treated at Melanoma Institute Australia (MIA) between 2001 and 2011 were analyzed. RESULTS: Interobserver agreement was excellent for American Joint Committee on Cancer (AJCC) T staging parameters: Breslow thickness (intraclass correlation coefficient [ICC] 0.984), mitotic rate (ICC 0.833), and ulceration (kappa statistic [κ] 0.823). All three of these important pathologic variables were included in 92.4 and 66.9% of review (MIA) and external (non-MIA) pathology reports, respectively. Completeness of MIA and non-MIA pathology reports for the three essential T-staging criteria increased significantly from 87.9 to 94.6% (χ(2) = 9.1, df = 1, P = 0.003) and from 53.2 to 74.3% (χ(2) = 35.0, df = 1, P < 0.001) over the 10-year study period. The AJCC N staging parameter of microsatellites was recorded in only 43% of non-MIA reports and demonstrated moderate concordance (κ = 0.560). CONCLUSIONS: Reproducibility and completeness of pathology reports for many important histopathologic features have improved in recent years. Nevertheless, the documentation of microsatellites remained poor in external pathology reports. To enhance the usefulness of the pathology report for the provision of optimal melanoma patient care, continued efforts to encourage pathologists to document its key features appear warranted.

Molecular alterations in metaplastic breast carcinoma.

Metaplastic carcinoma of the breast is a rare and heterogeneous subtype of breast carcinoma with a generally poor outcome, and few therapeutic options once disease recurs or progresses. Metaplastic carcinomas of the breast are usually of a larger size at diagnosis, with less frequent nodal metastasis compared with invasive ductal carcinoma no special type, and lack hormone and HER2 receptor expression. Recent research has revealed some potentially actionable genetic changes in a subset of these rare tumours. However, ongoing efforts to further characterise the genetic basis and the molecular alterations underlying the distinctive morphological and clinical characteristics of these tumours are needed in order to identify new targets for treatment. This review will describe the theories of pathogenesis of metaplastic breast carcinoma, and highlight genetic changes and potential therapeutic targets in this generally poor prognosis malignancy.

Recent insights into the molecular pathogenesis of mammary phyllodes tumours.

Phyllodes tumours (PTs) of the breast are true biphasic neoplasms within which interactions between the epithelium and stroma are critical for tumour development and progression. Despite numerous studies reporting the results of ancillary marker investigations in PTs, the current histological grading systems remain unreliable at predicting clinical outcome even when supplemented by these markers. As a consequence, there has been much interest in the prospect of using molecular/genetic techniques to develop a more robust "grading" system. This review focuses on recent cytogenetic and molecular studies investigating the pathogenesis of PTs and those correlating molecular findings with clinicopathological features of the tumours. Recent data highlight that intratumoural genetic heterogeneity is common in PTs and may account for the reported lack of correlation between histological grading and clinical behaviour. The entire spectrum of molecular aberrations in PTs are yet to be fully defined, however recent array-based studies using comparative genomic hybridisation have reported that copy number changes increase with the progression from benign PT to malignancy. Tumour recurrence and progression is likely to reflect the presence of under-recognised subclones. p(16INK4a) (CDKN2A) inactivation also appears to be important in PT pathogenesis. Further additional studies will be required to identify and validate new prognostic markers and therapeutic targets in order to improve the diagnosis, classification, prediction of outcome and management of patients with this rare neoplasm. Data generated from modern sequencing technologies are likely to provide new insights into the disease and assist in this endeavour.

Digital papillary adenocarcinoma: a tumour that should be considered in the differential diagnosis of neoplasms involving the digits.

BACKGROUND: Digital papillary adenocarcinoma (DPA) is a rare malignant adnexal tumour that occurs almost exclusively in a digital location. Only two case series have been reported previously. AIMS: To document the clinical and histopathological findings in a series of patients with DPA. RESULTS: Of the 8 patients identified, six were males and two were females, and they were aged between 32 and 76 years at diagnosis. The diagnosis was not clinically suspected in any of the patients, and the tumours were thought to be cysts in four cases. The tumours ranged from 7 mm to 40 mm in size, and were located on the right thumb (n=3), the right middle finger (n = 3) and the left index finger (n = 2). Each tumour exhibited a multinodular growth pattern, with a combination of solid and cystic architecture. Papillary projections were present in many of the cystic spaces. In three cases, some cystic spaces were lined by luminal columnar epithelial cells and underlying myoepithelial cells. Cytological atypia was generally mild and mitotic activity was low in most cases (median 2 mitoses per mm). Local tumour recurrence occurred in three patients, of whom one subsequently had a digital amputation. No metastases have been identified in the seven patients with known follow-up information. CONCLUSIONS: DPA is a rare tumour that should be included in the differential diagnosis of epithelial neoplasms in digital locations. Because of its tendency to recur locally, wide local excision should be performed as definitive initial treatment.