Dora Nagy

PURPOSE: The antibody-drug conjugate enfortumab vedotin (EV) releases a cytotoxic agent into tumor cells via binding to the membrane receptor NECTIN-4. EV was recently approved for patients with metastatic urothelial carcinoma (mUC) without prior assessment of the tumor receptor status as ubiquitous NECTIN-4 expression is assumed. Our objective was to determine the prevalence of membranous NECTIN-4 protein expression in primary tumors (PRIM) and patient-matched distant metastases (MET). EXPERIMENTAL DESIGN: Membranous NECTIN-4 protein expression was measured (H-score) by IHC in PRIM and corresponding MET (N = 137) and in a multicenter EV-treated cohort (N = 47). Progression-free survival (PFS) after initiation of EV treatment was assessed for the NECTIN-4-negative/weak (H-score 0-99) versus moderate/strong (H-score 100-300) subgroup. The specificity of the NECTIN-4 IHC staining protocol was validated by establishing CRISPR-Cas9-induced polyclonal NECTIN-4 knockouts. RESULTS: In our cohort, membranous NECTIN-4 expression significantly decreased during metastatic spread (Wilcoxon matched pairs P < 0.001; median H-score = 40; interquartile range, 0-140), with 39.4% of MET lacking membranous NECTIN-4 expression. In our multicenter EV cohort, absence or weak membranous NECTIN-4 expression (34.0% of the cohort) was associated with a significantly shortened PFS on EV (log-rank P < 0.001). CONCLUSIONS: Membranous NECTIN-4 expression is frequently decreased or absent in mUC tissue. Of note, the clinical benefit of EV strongly depends on membranous NECTIN-4 expression. Thus, our results are of highest clinical relevance and argue for a critical reconsideration of the current practice and suggest that the NECTIN-4 receptor status should be determined (ideally in a metastatic/progressive lesion) before initiation of EV. See related commentary by Aggen et al., p. 1377.

PURPOSE The anti-NECTIN4 antibody-drug conjugate enfortumab vedotin (EV) is approved for patients with metastatic urothelial cancer (mUC). However, durable benefit is only achieved in a small, yet uncharacterized patient subset. NECTIN4 is located on chromosome 1q23.3, and 1q23.3 gains represent frequent copy number variations (CNVs) in urothelial cancer. Here, we aimed to evaluate NECTIN4 amplifications as a genomic biomarker to predict EV response in patients with mUC. MATERIALS AND METHODS We established a NECTIN4-specific fluorescence in situ hybridization (FISH) assay to assess the predictive value of NECTIN4 CNVs in a multicenter EV-treated mUC patient cohort (mUC-EV, n = 108). CNVs were correlated with membranous NECTIN4 protein expression, EV treatment responses, and outcomes. We also assessed the prognostic value of NECTIN4 CNVs measured in metastatic biopsies of non–EV-treated mUC (mUC-non-EV, n = 103). Furthermore, we queried The Cancer Genome Atlas (TCGA) data sets (10,712 patients across 32 cancer types) for NECTIN4 CNVs. RESULTS NECTIN4 amplifications are frequent genomic events in muscle-invasive bladder cancer (TCGA bladder cancer data set: approximately 17%) and mUC (approximately 26% in our mUC cohorts). In mUC-EV, NECTIN4 amplification represents a stable genomic alteration during metastatic progression and associates with enhanced membranous NECTIN4 protein expression. Ninety-six percent (27 of 28) of patients with NECTIN4 amplifications demonstrated objective responses to EV compared with 32% (24 of 74) in the nonamplified subgroup ( P &lt; .001). In multivariable Cox analysis adjusted for age, sex, and Bellmunt risk factors, NECTIN4 amplifications led to a 92% risk reduction for death (hazard ratio, 0.08 [95% CI, 0.02 to 0.34]; P &lt; .001). In the mUC-non-EV, NECTIN4 amplifications were not associated with outcomes. TCGA Pan-Cancer analysis demonstrated that NECTIN4 amplifications occur frequently in other cancers, for example, in 5%-10% of breast and lung cancers. CONCLUSION NECTIN4 amplifications are genomic predictors of EV responses and long-term survival in patients with mUC.

673 Background: The anti-NECTIN4 antibody-drug conjugate (ADC) enfortumab vedotin (EV) is approved for patients with metastatic urothelial cancer (mUC). However, durable benefit is only achieved in a small, yet uncharacterized patient subset. NECTIN4 is located on chromosome 1q23.3, and 1q23.3 gains are frequent genomic events in mUC leading to NECTIN4 amplifications. Here, we aimed to evaluate the potential of NECTIN4 amplification as a genomic biomarker to predict EV response in patients with mUC. Methods: We established a NECTIN4-specific fluorescence in-situ hybridization (FISH) assay to assess NECTIN4 copy number variations (CNVs) in a multicenter EV-treated mUC patient cohort (UC-EV, N=77), and CNV data were correlated with membranous NECTIN4 protein expression (H-score) assessed via immunohistochemistry, EV treatment responses and outcomes. Next, we conducted a pan-cancer analysis of the The Cancer Genome Atlas (TCGA) datasets, comprising 10,712 patients across 32 cancer types, to investigate the relationship between NECTIN4 CNV, mRNA expression (RNAseq) and protein levels (RPPA) across entities. Results: In TCGA cohorts, NECTIN4 amplification occurs frequently across different solid cancer types, especially in 15-20% of bladder cancers (17% in TCGA-BLCA), as well as 5-10% in breast cancer (TCGA-BRCA) and lung adenocarcinoma (TCGA-LUAD). We confirmed the amplification frequency in our UC-EV cohort (18%). NECTIN4 amplification is significantly associated with both increased NECTIN4 mRNA expression (e.g., TCGA-BLCA, BRCA, LUAD) and membranous NECTIN4 protein expression (UC-EV), and represents a stable genomic alteration during metastatic progression. In the UC-EV cohort, all patients with NECTIN4 amplification (N=14) responded to EV. In multivariable Cox adjusted for age and sex, NECTIN4 amplifications led to a 93% risk reduction for death compared to a NECTIN4non-amplified status (HR=0.07, 95%-CI 0.01–0.53; P&lt;0.001). Conclusions: Our study highlights the value of NECTIN4 amplifications to predict EV responses in patients with mUC. NECTIN4 amplifications occur frequently in different cancer types and therefore have the potential to be a novel tumor-agnostic genomic biomarker that enables tailored NECTIN4-targeted therapies in various entities.

BACKGROUND: The shortage of pathologists in Germany, coupled with an aging workforce, requires innovative approaches to attract medical students to the field. Medical education must address different learning styles to ensure that all students are successful. METHODS: The pilot project "Practical Pathology" aims to enhance students' understanding of pathology by providing hands-on experience in macroscopic gross analysis through the use of tumor dummies built from scratch. RESULTS: An evaluation survey, completed by 63 participating students provided positive feedback on the course methodology, its relevance to understanding the pathology workflow, and its improvement over traditional teaching methods. The majority of students recognized the importance of hands-on training in medical education. Students with previous work experience rated the impact of the course on knowledge acquisition even more positively. CONCLUSION: The course improved students' understanding of pathological processes and potential sources of clinical-pathological misunderstanding. An increase in motivation for a potential career in the field of pathology was observed in a minority of students, although this exceeded the percentage of pathologists in the total medical workforce.

Inflammatory fibroid polyps (IFP) are rare and benign mesenchymal tumours of the gastrointestinal tract. They are submucosal spindle cell lesions with an eosinophilic-rich inflammatory infiltrate and mutations in the platelet-derived growth factor receptor alpha (PDGFRA) gene. In this report, we present the case of a 74-year-old female with a solid tumour of the kidney, which presented as a bland proliferation of spindle cells with thin-walled blood vessels and an inflammatory infiltrate with eosinophilic granulocytes. Immunohistochemistry revealed a positivity for vimentin and a weak staining for CD99 and CD34 in the spindle cells. Because of the morphological similarity to IFPs of the gastrointestinal tract, a molecular pathology analysis was performed. This identified an oncogenic mutation in exon 18 of the PDGFRA gene, which is characteristic for inflammatory fibroid polyps of the gastrointestinal tract. To the best of our knowledge, this is the first case of an IFP in the urogenital tract.