Background Breast cancer in young adults has been implicated with a worse outcome. Analyses of genomic traits associated with age have been heterogenous, likely because of an incomplete accounting for underlying molecular subtypes. We aimed to resolve whether triple-negative breast cancer (TNBC) in younger versus older patients represent similar or different molecular diseases in the context of genetic and transcriptional subtypes and immune cell infiltration. Patients and methods In total, 237 patients from a reported population-based south Swedish TNBC cohort profiled by RNA sequencing and whole-genome sequencing (WGS) were included. Patients were binned in 10-year intervals. Complimentary PD-L1 and CD20 immunohistochemistry and estimation of tumor-infiltrating lymphocytes (TILs) were performed. Cases were analyzed for differences in patient outcome, genomic, transcriptional, and immune landscape features versus age at diagnosis. Additionally, 560 public WGS breast cancer profiles were used for validation. Results Median age at diagnosis was 62 years (range 26–91). Age was not associated with invasive disease-free survival or overall survival after adjuvant chemotherapy. Among the BRCA1-deficient cases (82/237), 90% were diagnosed before the age of 70 and were predominantly of the basal-like subtype. In the full TNBC cohort, reported associations of patient age with changes in Ki67 expression, PIK3CA mutations, and a luminal androgen receptor subtype were confirmed. Within DNA repair deficiency or gene expression defined molecular subgroups, age-related alterations in, e.g., overall gene expression, immune cell marker gene expression, genetic mutational and rearrangement signatures, amount of copy number alterations, and tumor mutational burden did, however, not appear distinct. Similar non-significant associations for genetic alterations with age were obtained for other breast cancer subgroups in public WGS data. Consistent with age-related immunosenescence, TIL counts decreased linearly with patient age across different genetic TNBC subtypes. Conclusions Age-related alterations in TNBC, as well as breast cancer in general, need to be viewed in the context of underlying genomic phenotypes. Based on this notion, age at diagnosis alone does not appear to provide an additional layer of biological complexity above that of proposed genetic and transcriptional phenotypes of TNBC. Consequently, treatment decisions should be less influenced by age and more driven by tumor biology.

Prevalences of diverticulum and low-grade mucinous neoplasm (LGMN) were reported as 0.04-2% and less than 1% in the appendix. In this study, the frequency of diverticulum in LGMN cases, the relationship between diverticula and periappendicular mucin, and the possible role of diverticula in pseudomyxoma peritonei pathogenesis were researched. Through systematic review and targeted search, 38 LGMN and 96 diverticula were identified, frequencies and relationship between diverticulum and LGMN were analysed. Diverticulum and LGMN were determined in 4.8% and 1.04%, respectively, of 1922 appendectomy materials specifically grossed by the same pathologist. The prevalence of diverticulum was higher in our study than literature. The difference may be due to detailed macroscopic examination. Diverticulum was detected in 60% of LGMN cases. The rate of diverticulum was found to be significantly higher in LGMNs than non-neoplastic diverticulum (p < 0.001). Periappendicular mucin deposition was significantly more frequent in LGMN cases with diverticulum than in other groups (p < 0.05). Follow-ups were available in 18 patients; none of them had mucin deposition in the peritoneal cavity. We detected that periappendicular mucin was highly associated with diverticula in LGMN cases. Periappendicular acellular mucin deposition may not give rise to pseudomyxoma peritonei. We may think that mucin could move out of the appendix through the diverticulum rather than neoplastic spread in some of these cases.

Ki67 is an important biomarker with prognostic and potential predictive value in breast cancer. However, the lack of standardization hinders its clinical applicability. In this study, we aimed to investigate the reproducibility among pathologists following the guidelines of the International Ki67 in Breast Cancer Working Group (IKWG) for Ki67 scoring and to evaluate the prognostic potential of this platform in an independent cohort. Four algorithms were independently built by four pathologists based on our study cohort using an open-source digital image analysis (DIA) platform (QuPath) following the detailed guideline of the IKWG. The algorithms were applied on an ER+ breast cancer study cohort of 157 patients with 15 years of follow-up. The reference Ki67 score was obtained by a DIA algorithm trained on a subset of the study cohort. Intraclass correlation coefficient (ICC) was used to measure reproducibility. High interobserver reliability was reached with an ICC of 0.938 (CI: 0.920–0.952) among the algorithms and the reference standard. Comparing each machine-read score against relapse-free survival, the hazard ratios were similar (2.593–4.165) and showed independent prognostic potential (p ≤ 0.018, for all comparisons). In conclusion, we demonstrate high reproducibility and independent prognostic potential using the IKWG DIA instructions to score Ki67 in breast cancer. A prospective study is needed to assess the clinical utility of the IKWG DIA Ki67 instructions.

Pseudoangiomatous stromal hyperplasia (PASH) is a benign proliferative entity of mammary stroma. It is generally found as an incidental finding. It may rarely present as a palpable nodule. Three patients, who were 29, 45, and 58 years of age, were referred to our clinic with nodule and pain in the breast. The physical examinations and ultrasound findings of all three patients were consistent with fibroadenoma. Core biopsies were performed and reported as "benign breast parenchyma including stromal fibrosis." PASH areas were noted in one case. The excision specimens were observed as solid nodular masses with smooth external surfaces and white in colour. Microscopically, well-demarcated hyalinized stroma, including slit-like pseudovascular spaces lined by bland spindle cells, was observed. Immunohistochemically, these cells showed positive staining for CD34 and negative staining for CD31. Nodule-forming PASH mostly confuses with fibroadenoma with respect to clinical examination and radiological findings. Definite diagnosis requires histopathological verification. Differential diagnosis should be made with low grade angiosarcoma, fibroepithelial tumors, and myofibroblastoma.

Urothelial carcinoma with abundant myxoid stroma is a newly-described and extremely rare entity. Since only very few cases have been reported, there is no consensus on its nomenclature. Microscopic examination revealed invasive urothelial carcinoma with widespread low-grade noninvasive areas. There were focal invasive areas in the neighborhood of the renal parenchyma. Malignant urothelial tumor/cell groups localized in the stroma had abundant myxoid/mucinous background in the invasive areas. The cytoplasm of the tumoral cells was more eosinophilic in these areas and the cells formed small groups and cords. Histochemically, PAS and Alcian Blue were positive in the cytoplasm of the tumoral cells and in the stroma while negative in the non-mucinous areas. Immunohistochemically, the tumoral cells of the mucinous invasive areas diffusely expressed MUC1 and MUC2. We discuss the origin of the mucinous/myxoid stroma, the tumor’s nature and its nomenclature with histochemical and immunohistochemical features.

SUMMARYWe present two cases of pleomorphic adenoma, one that developed in the breast parenchyma and the other in the breast skin, with their histopathological differential diagnosis. BACKGROUND

Emerging data indicate that genomic alterations can shape immune cell composition in early breast cancer. However, there is a need for complementary imaging and sequencing methods for the quantitative assessment of combined somatic copy number alteration (SCNA) and immune profiling in pathological samples. Here, we tested the feasibility of three approaches—CUTseq, for high-throughput low-input SCNA profiling, multiplexed fluorescent immunohistochemistry (mfIHC) and digital-image analysis (DIA) for quantitative immuno-profiling- in archival formalin-fixed paraffin-embedded (FFPE) tissue samples from patients enrolled in the randomized SBG-2004-1 phase II trial. CUTseq was able to reproducibly identify amplification and deletion events with a resolution of 100 kb using only 6 ng of DNA extracted from FFPE tissue and pooling together 77 samples into the same sequencing library. In the same samples, mfIHC revealed that CD4 + T-cells and CD68 + macrophages were the most abundant immune cells and they mostly expressed PD-L1 and PD-1. Combined analysis showed that the SCNA burden was inversely associated with lymphocytic infiltration. Our results set the basis for further applications of CUTseq, mfIHC and DIA to larger cohorts of early breast cancer patients.

PASH lesions occurred with a frequency of 9% in CNBS and were mostly in association with benign breast lesions in premenopausal women. CCC was determined as an accompanying epithelial lesion within or near PASH areas. No obvious immunopositivity compatible with endothelial cell differentiation was revealed.