Integrating NECTIN4 Amplification with Membranous NECTIN4 Expression to Develop a Scoring System for Predicting Enfortumab Vedotin Response in Urothelial Carcinoma

Clinical Cancer Research, pp. OF1–OF14

Abstract

PURPOSE: Enfortumab vedotin (EV) is standard therapy for metastatic urothelial carcinoma (mUC), yet the predictive relevance of NECTIN4 expression-especially membranous versus cytoplasmic-remains unclear. Here, we sought to extend previous findings on NECTIN4 gene amplification in parallel with a systematic subcellular evaluation of NECTIN4 expression. EXPERIMENTAL DESIGN: We retrospectively analyzed 179 EV-treated mUC patients. NECTIN4 amplification was assessed by FISH and NECTIN4 protein levels by IHC. A four-tier membranous scoring algorithm (0,1+,2+,3+) adapted from CAP HER2 gastric guidelines was benchmarked against H-score. We integrated amplification status with membranous staining to refine predictive stratification and compared associations with objective response rate (ORR) to EV-301 data. RESULTS: Combining membranous and cytoplasmic compartments resulted in a median composite H-score of 260 (78.2% ≥150), closely matching NECTIN4 expression prevalence reported in EV-301 (median 250; 82.6% ≥ 150). A ≥150 cut-off enriched for EV responders in both cohorts; in EV-301 with ORR of 45.8% vs. 20% (P = 0.001). High membranous expression based on the scoring (2+/3+) predicted response (ORR 55.1% vs. 25.5%; P < 0.001), with longer PFS (7.1 vs. 2.9 months; HR 0.45) and OS (12.3 vs. 6.9 months; HR 0.57), whereas cytoplasmic expression lacked predictive value. NECTIN4-amplified tumors showed particularly favorable outcomes (PFS 12.2 months; OS 30.1 months). An integrated three-tier model-amplified, non-amplified/high-membranous, and non-amplified/low-membranous-yielded ORRs of 77.2%, 42.9%, and 26.1% and separated survival outcomes. CONCLUSIONS: Our NECTIN4 scoring system integrating NECTIN4 amplification with membranous NECTIN4 expression accurately predicts outcomes, supporting combined genomic and membranous assessment as complementary biomarkers for optimizing EV selection.

Authors 43

1. Niklas Klümper corresponding

   University Hospital Bonn

   Affiliation as printed

   2Institute of Experimental Oncology, University Medical Center Bonn (UKB), Bonn, Germany

   3Center for Integrated Oncology Aachen/Bonn/Cologne/Düsseldorf (CIO-ABCD), Bonn, Germany

   4BRIDGE-Consortium Germany e.V., Mannheim, Germany

   University Hospital Bonn 1 Department of Urology and Pediatric Urology, , Bonn, Germany

2. Thomas Büttner

   University Hospital Bonn

   Affiliation as printed

   3Center for Integrated Oncology Aachen/Bonn/Cologne/Düsseldorf (CIO-ABCD), Bonn, Germany

   4BRIDGE-Consortium Germany e.V., Mannheim, Germany

   University Hospital Bonn 1 Department of Urology and Pediatric Urology, , Bonn, Germany

3. Sebastian Rauch

   Friedrich-Alexander-Universität Erlangen-Nürnberg · Universitätsklinikum Erlangen · Helmholtz Institute Erlangen-Nürnberg · Comprehensive Cancer Center Erlangen

   Affiliation as printed

   5Institute of Pathology, University Hospital Erlangen, Friedrich-Alexander-Universität Erlangen-Nürnberg (FAU), Erlangen, Germany

   7Bavarian Center for Cancer Research (Bayerisches Zentrum für Krebsforschung, BZKF), Erlangen, Germany

   Friedrich-Alexander-Universität Erlangen-Nürnberg 6 Comprehensive Cancer Center EMN, University Hospital Erlangen, , Erlangen, Germany

4. Stefanie Zschäbitz

   Heidelberg University · University Hospital Heidelberg · National Center for Tumor Diseases

   Affiliation as printed

   8Department of Medical Oncology, National Center for Tumor Disease (NCT), University Hospital, Heidelberg, Germany

5. Florian Roghmann

   Universitätsklinik Marien Hospital Herne · Ruhr University Bochum

   Affiliation as printed

   4BRIDGE-Consortium Germany e.V., Mannheim, Germany

   9Department of Urology, Marien Hospital, Ruhr-University Bochum, Herne, Germany

6. Christian Bolenz

   Universität Ulm · University Hospital Ulm

   Affiliation as printed

   4BRIDGE-Consortium Germany e.V., Mannheim, Germany

   University Hospital Ulm 10 Department of Urology and Pediatric Urology, , University of Ulm, Ulm, Germany

7. Fabienne Lange

   Friedrich-Alexander-Universität Erlangen-Nürnberg · Universitätsklinikum Erlangen · Helmholtz Institute Erlangen-Nürnberg · Comprehensive Cancer Center Erlangen

   Affiliation as printed

   5Institute of Pathology, University Hospital Erlangen, Friedrich-Alexander-Universität Erlangen-Nürnberg (FAU), Erlangen, Germany

   7Bavarian Center for Cancer Research (Bayerisches Zentrum für Krebsforschung, BZKF), Erlangen, Germany

   Friedrich-Alexander-Universität Erlangen-Nürnberg 6 Comprehensive Cancer Center EMN, University Hospital Erlangen, , Erlangen, Germany

8. Patricia C. Rausch

   Universitätsklinik Marien Hospital Herne · Ruhr University Bochum

   Affiliation as printed

   4BRIDGE-Consortium Germany e.V., Mannheim, Germany

   9Department of Urology, Marien Hospital, Ruhr-University Bochum, Herne, Germany

9. Friedemann Zengerling

   Universität Ulm · University Hospital Ulm

   Affiliation as printed

   4BRIDGE-Consortium Germany e.V., Mannheim, Germany

   University Hospital Ulm 10 Department of Urology and Pediatric Urology, , University of Ulm, Ulm, Germany

10. Nadine Therese Gaisa

    Universität Ulm · University Hospital Ulm

    Affiliation as printed

    4BRIDGE-Consortium Germany e.V., Mannheim, Germany

    University Hospital Ulm 11 Institute of Pathology, , University of Ulm, Ulm, Germany

11. Oliver Hahn

    University of Würzburg · Universitätsklinikum Würzburg

    Affiliation as printed

    12Department of Urology and Pediatric Urology, Julius Maximilians University Medical Center of Würzburg, Würzburg, Germany

12. Constantin Schwab

    Heidelberg University

    Affiliation as printed

    13Institute of Pathology, University of Heidelberg, Heidelberg, Germany

13. Anna Katrin Scherping

    University Hospital Bonn

    Affiliation as printed

    3Center for Integrated Oncology Aachen/Bonn/Cologne/Düsseldorf (CIO-ABCD), Bonn, Germany

    University Hospital Bonn 14 Institute of Pathology, , Bonn, Germany

14. Marieta Ioana Toma

    University Hospital Bonn

    Affiliation as printed

    3Center for Integrated Oncology Aachen/Bonn/Cologne/Düsseldorf (CIO-ABCD), Bonn, Germany

    University Hospital Bonn 14 Institute of Pathology, , Bonn, Germany

15. Hendrik Heers Aachen 15Department of Urology

    Universitätsklinikum Aachen

    Affiliation as printed

    15Department of Urology, University Hospital Aachen, Aachen, Germany

16. Philipp Ivanyi

    Medizinische Hochschule Hannover

    Affiliation as printed

    16Department of Hemostaseology, Oncology and Stem Cell Transplantation, Medical University Hannover, Hannover, Germany

17. Günter Niegisch

    Düsseldorf University Hospital

    Affiliation as printed

    17Department of Urology, University Hospital Düsseldorf, Düsseldorf, Germany

    3Center for Integrated Oncology Aachen/Bonn/Cologne/Düsseldorf (CIO-ABCD), Bonn, Germany

18. Camilla Marisa Grunewald

    Düsseldorf University Hospital

    Affiliation as printed

    17Department of Urology, University Hospital Düsseldorf, Düsseldorf, Germany

19. Christopher Darr

    Essen University Hospital

    Affiliation as printed

    18Department of Urology, University Hospital Essen, Essen, Germany

20. Pia Paffenholz

    University Hospital Cologne

    Affiliation as printed

    University Hospital Cologne 19 Department of Urology, , Cologne, Germany

21. Richard Weiten

    University Hospital Cologne

    Affiliation as printed

    University Hospital Cologne 19 Department of Urology, , Cologne, Germany

22. Susanne Fuessel

    University Hospital Carl Gustav Carus

    Affiliation as printed

    20Department of Urology, University Hospital Dresden, Dresden, Germany

23. Christian Thomas

    University Hospital Carl Gustav Carus

    Affiliation as printed

    20Department of Urology, University Hospital Dresden, Dresden, Germany

24. Marcus Sondermann

    University Hospital Carl Gustav Carus

    Affiliation as printed

    20Department of Urology, University Hospital Dresden, Dresden, Germany

25. Ulrich Sommer

    University Hospital Carl Gustav Carus

    Affiliation as printed

    20Department of Urology, University Hospital Dresden, Dresden, Germany

26. Kerstin Junker

    Saarland University

    Affiliation as printed

    Saarland University 21 Department of Urology and Pediatric Urology, , Homburg, Germany

27. Katrin Schlack

    University Hospital Münster

    Affiliation as printed

    22Department of Urology, University Hospital Münster, Münster, Germany

28. Jozefina Casuscelli

    Ludwig-Maximilians-Universität München

    Affiliation as printed

    23Department of Urology, University Hospital, Ludwig Maximilian University of Munich, Munich, Germany

29. Can Daniel Aydogdu

    Ludwig-Maximilians-Universität München

    Affiliation as printed

    23Department of Urology, University Hospital, Ludwig Maximilian University of Munich, Munich, Germany

30. Dora Niedersuess‐Beke
    Affiliation as printed

    24Department of Internal Medicine I., Centre for Oncology and Haematology, Vienna Healthcare Group, Vienna, Austria

31. Steffen Rausch

    University of Tübingen

    Affiliation as printed

    25Department of Urology, Eberhard Karls University, Tübingen, Germany

32. Christoph Kuppe Aachen Division of Nephrology and Clinical Immunology

    RWTH Aachen University

    Affiliation as printed

    RWTH Aachen University 26 Medical Faculty, Division of Nephrology and Clinical Immunology, , Aachen, Germany

33. Joshua J. Meeks
    Affiliation as printed

    27Department of Urology, Feinberg School of Medicine, Chicago, Illinois

34. Bonnie Choy
    Affiliation as printed

    28Department of Pathology, Feinberg School of Medicine, Chicago, Illinois

35. Sarmad Sadeghi

    University of Southern California · USC Norris Comprehensive Cancer Center

    Affiliation as printed

    USC - University of Southern California 29 Medicine Department, , Norris Comprehensive Cancer Center, Los Angeles, California

36. Parkash S. Gill

    University of Southern California · USC Norris Comprehensive Cancer Center

    Affiliation as printed

    USC - University of Southern California 29 Medicine Department, , Norris Comprehensive Cancer Center, Los Angeles, California

37. Manuel Ritter

    University Hospital Bonn

    Affiliation as printed

    3Center for Integrated Oncology Aachen/Bonn/Cologne/Düsseldorf (CIO-ABCD), Bonn, Germany

    4BRIDGE-Consortium Germany e.V., Mannheim, Germany

    University Hospital Bonn 1 Department of Urology and Pediatric Urology, , Bonn, Germany

38. Johannes Brägelmann

    University of Bonn · University Hospital Bonn

    Affiliation as printed

    30Medical Clinic III for Oncology, Hematology, Immune-Oncology and Rheumatology, University Medical Center Bonn (UKB), Bonn, Germany

    3Center for Integrated Oncology Aachen/Bonn/Cologne/Düsseldorf (CIO-ABCD), Bonn, Germany

39. Michael Hölzel

    University Hospital Bonn

    Affiliation as printed

    2Institute of Experimental Oncology, University Medical Center Bonn (UKB), Bonn, Germany

    3Center for Integrated Oncology Aachen/Bonn/Cologne/Düsseldorf (CIO-ABCD), Bonn, Germany

40. Arndt Hartmann

    Friedrich-Alexander-Universität Erlangen-Nürnberg · Universitätsklinikum Erlangen · Helmholtz Institute Erlangen-Nürnberg · Comprehensive Cancer Center Erlangen

    Affiliation as printed

    4BRIDGE-Consortium Germany e.V., Mannheim, Germany

    5Institute of Pathology, University Hospital Erlangen, Friedrich-Alexander-Universität Erlangen-Nürnberg (FAU), Erlangen, Germany

    7Bavarian Center for Cancer Research (Bayerisches Zentrum für Krebsforschung, BZKF), Erlangen, Germany

    Friedrich-Alexander-Universität Erlangen-Nürnberg 6 Comprehensive Cancer Center EMN, University Hospital Erlangen, , Erlangen, Germany

41. Viktor Grünwald

    Essen University Hospital

    Affiliation as printed

    31Clinic for Internal Medicine (Tumor Research) and Clinic for Urology, Interdisciplinary Genitourinary Oncology at the West-German Cancer Center, Essen University Hospital, Essen, Germany

42. Jonas Saal

    University of Bonn · University Hospital Bonn

    Affiliation as printed

    2Institute of Experimental Oncology, University Medical Center Bonn (UKB), Bonn, Germany

    30Medical Clinic III for Oncology, Hematology, Immune-Oncology and Rheumatology, University Medical Center Bonn (UKB), Bonn, Germany

    3Center for Integrated Oncology Aachen/Bonn/Cologne/Düsseldorf (CIO-ABCD), Bonn, Germany

43. Markus Eckstein

    Friedrich-Alexander-Universität Erlangen-Nürnberg · Universitätsklinikum Erlangen · Helmholtz Institute Erlangen-Nürnberg · Comprehensive Cancer Center Erlangen

    Affiliation as printed

    4BRIDGE-Consortium Germany e.V., Mannheim, Germany

    5Institute of Pathology, University Hospital Erlangen, Friedrich-Alexander-Universität Erlangen-Nürnberg (FAU), Erlangen, Germany

    7Bavarian Center for Cancer Research (Bayerisches Zentrum für Krebsforschung, BZKF), Erlangen, Germany

    Friedrich-Alexander-Universität Erlangen-Nürnberg 6 Comprehensive Cancer Center EMN, University Hospital Erlangen, , Erlangen, Germany