PD-L1 targeting and subclonal immune escape mediated by PD-L1 mutations in metastatic colorectal cancer
Background In patients with microsatellite stable (MSS) metastatic colorectal cancer (mCRC), immune checkpoint blockade is ineffective, and combinatorial approaches enhancing immunogenicity need exploration.Methods We treated 43 patients with predominantly microsatellite stable RAS/BRAF wild-type mC...
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BMJ Publishing Group
2021-07-01
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| Series: | Journal for ImmunoTherapy of Cancer |
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| author | Thomas J Ettrich Barbara Seliger Chiara Massa Alexander Stein Claudia Wickenhauser Axel Hinke Uwe Pelzer Carsten Bokemeyer Marcus Bauer Susanna Hegewisch-Becker Donjete Simnica Christoph Schultheiß Rebekka Scholz Joseph Tintelnot Eray Gökkurt Lisa von Wenserski Edith Willscher Lisa Paschold Markus Sauer Sylvie Lorenzen Jorge Riera-Knorrenschild Reinhard Depenbusch Steffen Dörfel Salah-Eddin Al-Batran Meinolf Karthaus Lisa Waberer Mascha Binder |
| author_facet | Thomas J Ettrich Barbara Seliger Chiara Massa Alexander Stein Claudia Wickenhauser Axel Hinke Uwe Pelzer Carsten Bokemeyer Marcus Bauer Susanna Hegewisch-Becker Donjete Simnica Christoph Schultheiß Rebekka Scholz Joseph Tintelnot Eray Gökkurt Lisa von Wenserski Edith Willscher Lisa Paschold Markus Sauer Sylvie Lorenzen Jorge Riera-Knorrenschild Reinhard Depenbusch Steffen Dörfel Salah-Eddin Al-Batran Meinolf Karthaus Lisa Waberer Mascha Binder |
| author_sort | Thomas J Ettrich |
| collection | DOAJ |
| description | Background In patients with microsatellite stable (MSS) metastatic colorectal cancer (mCRC), immune checkpoint blockade is ineffective, and combinatorial approaches enhancing immunogenicity need exploration.Methods We treated 43 patients with predominantly microsatellite stable RAS/BRAF wild-type mCRC on a phase II trial combining chemotherapy with the epidermal growth factor receptor antibody cetuximab and the programmed cell death ligand 1 (PD-L1) antibody avelumab. We performed next-generation gene panel sequencing for mutational typing of tumors and liquid biopsy monitoring as well as digital droplet PCR to confirm individual mutations. Translational analyses included tissue immunohistochemistry, multispectral imaging and repertoire sequencing of tumor-infiltrating T cells. Detected PD-L1 mutations were mechanistically validated in CRISPR/Cas9-generated cell models using qRT-PCR, immunoblotting, flow cytometry, complement-dependent cytotoxicity assay, antibody-dependent cytotoxicity by natural killer cell degranulation assay and LDH release assay as well as live cell imaging of T cell mediated tumor cell killing.Results Circulating tumor DNA showed rapid clearance in the majority of patients mirroring a high rate of early tumor shrinkage. In 3 of 13 patients expressing the high-affinity Fcγ receptor 3a (FcγR3a), tumor subclones with PD-L1 mutations were selected that led to loss of tumor PD-L1 by nonsense-mediated RNA decay in PD-L1 K162fs and protein degradation in PD-L1 L88S. As a consequence, avelumab binding and antibody-dependent cytotoxicity were impaired, while T cell killing of these variant clones was increased. Interestingly, PD-L1 mutant subclones showed slow selection dynamics reversing on avelumab withdrawal and patients with such subclones had above-average treatment benefit. This suggested that the PD-L1 mutations mediated resistance to direct antitumor effects of avelumab, while at the same time loss of PD-L1 reduced biological fitness by enhanced T cell killing limiting subclonal expansion.Conclusion The addition of avelumab to standard treatment appeared feasible and safe. PD-L1 mutations mediate subclonal immune escape to avelumab in some patients with mCRC expressing high-affinity FcγR3a, which may be a subset experiencing most selective pressure. Future trials evaluating the addition of avelumab to standard treatment in MSS mCRC are warranted especially in this patient subpopulation.Trial registration number NCT03174405. |
| format | Article |
| id | doaj-art-5d88a304ea0e44abaa1cca6a22c6374b |
| institution | Kabale University |
| issn | 2051-1426 |
| language | English |
| publishDate | 2021-07-01 |
| publisher | BMJ Publishing Group |
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| series | Journal for ImmunoTherapy of Cancer |
| spelling | doaj-art-5d88a304ea0e44abaa1cca6a22c6374b2024-11-08T12:10:08ZengBMJ Publishing GroupJournal for ImmunoTherapy of Cancer2051-14262021-07-019710.1136/jitc-2021-002844PD-L1 targeting and subclonal immune escape mediated by PD-L1 mutations in metastatic colorectal cancerThomas J Ettrich0Barbara Seliger1Chiara Massa2Alexander Stein3Claudia Wickenhauser4Axel Hinke5Uwe Pelzer6Carsten Bokemeyer7Marcus Bauer8Susanna Hegewisch-Becker9Donjete Simnica10Christoph Schultheiß11Rebekka Scholz12Joseph Tintelnot13Eray Gökkurt14Lisa von Wenserski15Edith Willscher16Lisa Paschold17Markus Sauer18Sylvie Lorenzen19Jorge Riera-Knorrenschild20Reinhard Depenbusch21Steffen Dörfel22Salah-Eddin Al-Batran23Meinolf Karthaus24Lisa Waberer25Mascha Binder26Department of Internal Medicine I, University Hospital Ulm, Ulm, Baden-Württemberg, GermanyInstitute of Medical Immunology, Martin-Luther-Universitat Halle-Wittenberg, Halle, GermanyInstitute of Medical Immunology, Martin-Luther-Universitat Halle-Wittenberg, Halle, GermanyHematology-Oncology Practice Eppendorf (HOPE), Hamburg, GermanyInstitute of Pathology, University Hospital Halle, Halle, GermanyClinical Cancer Research Consulting (CCRC), Düsseldorf, GermanyDepartment of Hematology, Oncology and Tumorimmunology, Charite Universitatsmedizin Berlin, Berlin, GermanyDepartment of Internal Medicine II (Oncology/Hematology/BMT/Pneumology), University Medical Center Hamburg-Eppendorf, Hamburg, GermanyInstitute of Pathology, Martin Luther University Halle Wittenberg, Halle, Sachsen-Anhalt, GermanyHämatologisch-Onkologische Praxis Eppendorf, Hamburg, GermanyDepartment of Internal Medicine IV – Oncology/Hematology, Martin-Luther-Universitat Halle-Wittenberg, Halle, Sachsen-Anhalt, GermanyDepartment of Internal Medicine IV – Oncology/Hematology, Martin-Luther-Universitat Halle-Wittenberg, Halle, Sachsen-Anhalt, GermanyDepartment of Internal Medicine IV – Oncology/Hematology, Martin-Luther-Universitat Halle-Wittenberg, Halle, Sachsen-Anhalt, GermanyUniversity Cancer Center Hamburg (UCCH), University Medical Center Hamburg-Eppendorf, Hamburg, GermanyHämatologisch-Onkologische Praxis Eppendorf, Hamburg, GermanyDepartment of Internal Medicine IV – Oncology/Hematology, Martin-Luther-Universitat Halle-Wittenberg, Halle, Sachsen-Anhalt, GermanyDepartment of Internal Medicine IV – Oncology/Hematology, Martin-Luther-Universitat Halle-Wittenberg, Halle, Sachsen-Anhalt, GermanyInternal Medicine IV, Martin-Luther-University Halle-Wittenberg, Halle, GermanyDepartment of Oncology and Hematology, Bone Marrow Transplantation with Section Pneumology, University Medical Center Hamburg-Eppendorf, Hamburg, GermanyDepartment of Internal Medicine III (Haematology/Medical Oncology), Technical University of Munich Hospital Rechts der Isar, Munchen, Bayern, GermanyUniversity Hospital of Giessen and Marburg Campus Marburg, Marburg, Hessen, GermanyPrivate Practice Onkodoc GmbH Gütersloh, Gütersloh, Nordrhein-Westfalen, GermanyPrivate Practice Onkozentrum Dresden, Dresden, Sachsen, GermanyInstitute of Clinical Cancer Research IKF at Northwest hospital, Frankfurt, Hessen, GermanyDepartment of Hematology and Oncology, Munich Hospital Neuperlach, Munchen, Bayern, GermanyIKF Klinische Krebsforschung GmbH at Krankenhaus Nordwest, Frankfurt, Hessen, GermanyDivision of Medical Oncology, University Hospital Basel, Basel, SwitzerlandBackground In patients with microsatellite stable (MSS) metastatic colorectal cancer (mCRC), immune checkpoint blockade is ineffective, and combinatorial approaches enhancing immunogenicity need exploration.Methods We treated 43 patients with predominantly microsatellite stable RAS/BRAF wild-type mCRC on a phase II trial combining chemotherapy with the epidermal growth factor receptor antibody cetuximab and the programmed cell death ligand 1 (PD-L1) antibody avelumab. We performed next-generation gene panel sequencing for mutational typing of tumors and liquid biopsy monitoring as well as digital droplet PCR to confirm individual mutations. Translational analyses included tissue immunohistochemistry, multispectral imaging and repertoire sequencing of tumor-infiltrating T cells. Detected PD-L1 mutations were mechanistically validated in CRISPR/Cas9-generated cell models using qRT-PCR, immunoblotting, flow cytometry, complement-dependent cytotoxicity assay, antibody-dependent cytotoxicity by natural killer cell degranulation assay and LDH release assay as well as live cell imaging of T cell mediated tumor cell killing.Results Circulating tumor DNA showed rapid clearance in the majority of patients mirroring a high rate of early tumor shrinkage. In 3 of 13 patients expressing the high-affinity Fcγ receptor 3a (FcγR3a), tumor subclones with PD-L1 mutations were selected that led to loss of tumor PD-L1 by nonsense-mediated RNA decay in PD-L1 K162fs and protein degradation in PD-L1 L88S. As a consequence, avelumab binding and antibody-dependent cytotoxicity were impaired, while T cell killing of these variant clones was increased. Interestingly, PD-L1 mutant subclones showed slow selection dynamics reversing on avelumab withdrawal and patients with such subclones had above-average treatment benefit. This suggested that the PD-L1 mutations mediated resistance to direct antitumor effects of avelumab, while at the same time loss of PD-L1 reduced biological fitness by enhanced T cell killing limiting subclonal expansion.Conclusion The addition of avelumab to standard treatment appeared feasible and safe. PD-L1 mutations mediate subclonal immune escape to avelumab in some patients with mCRC expressing high-affinity FcγR3a, which may be a subset experiencing most selective pressure. Future trials evaluating the addition of avelumab to standard treatment in MSS mCRC are warranted especially in this patient subpopulation.Trial registration number NCT03174405.https://jitc.bmj.com/content/9/7/e002844.full |
| spellingShingle | Thomas J Ettrich Barbara Seliger Chiara Massa Alexander Stein Claudia Wickenhauser Axel Hinke Uwe Pelzer Carsten Bokemeyer Marcus Bauer Susanna Hegewisch-Becker Donjete Simnica Christoph Schultheiß Rebekka Scholz Joseph Tintelnot Eray Gökkurt Lisa von Wenserski Edith Willscher Lisa Paschold Markus Sauer Sylvie Lorenzen Jorge Riera-Knorrenschild Reinhard Depenbusch Steffen Dörfel Salah-Eddin Al-Batran Meinolf Karthaus Lisa Waberer Mascha Binder PD-L1 targeting and subclonal immune escape mediated by PD-L1 mutations in metastatic colorectal cancer Journal for ImmunoTherapy of Cancer |
| title | PD-L1 targeting and subclonal immune escape mediated by PD-L1 mutations in metastatic colorectal cancer |
| title_full | PD-L1 targeting and subclonal immune escape mediated by PD-L1 mutations in metastatic colorectal cancer |
| title_fullStr | PD-L1 targeting and subclonal immune escape mediated by PD-L1 mutations in metastatic colorectal cancer |
| title_full_unstemmed | PD-L1 targeting and subclonal immune escape mediated by PD-L1 mutations in metastatic colorectal cancer |
| title_short | PD-L1 targeting and subclonal immune escape mediated by PD-L1 mutations in metastatic colorectal cancer |
| title_sort | pd l1 targeting and subclonal immune escape mediated by pd l1 mutations in metastatic colorectal cancer |
| url | https://jitc.bmj.com/content/9/7/e002844.full |
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