PD-L1 targeting and subclonal immune escape mediated by PD-L1 mutations in metastatic colorectal cancer
Alexander Stein1,2, Donjete Simnica3, Christoph Schultheiß3
1Hämatologisch-Onkologische Praxis Eppendorf, Hamburg, Germany.
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.
Insights
In microsatellite stable metastatic colorectal cancer (mCRC), PD-L1 mutations can cause immune escape from avelumab treatment. However, these mutations may also enhance T cell killing, suggesting a complex role in treatment benefit.
Area of Science:
- Oncology
- Immunotherapy
- Colorectal Cancer Research
Background:
- Immune checkpoint blockade is ineffective in microsatellite stable (MSS) metastatic colorectal cancer (mCRC).
- Combinatorial approaches are needed to enhance immunogenicity in MSS mCRC.
Purpose of the Study:
- To evaluate the safety and efficacy of combining chemotherapy with cetuximab and avelumab in MSS mCRC patients.
- To investigate the mechanisms of potential resistance and response to this combination therapy.
Main Methods:
- A phase II trial combining chemotherapy, cetuximab, and avelumab in 43 MSS mCRC patients.
- Next-generation sequencing, liquid biopsy, digital droplet PCR, immunohistochemistry, and multispectral imaging for translational analyses.
- Mechanistic validation of PD-L1 mutations in CRISPR/Cas9 models.
Main Results:
- The combination therapy was feasible and safe, with early tumor shrinkage observed.
- PD-L1 mutations leading to loss of PD-L1 expression were selected in some patients with high-affinity FcγR3a.
- These PD-L1 mutations impaired avelumab binding but increased T cell killing, with complex effects on treatment benefit.
Conclusions:
- PD-L1 mutations can mediate subclonal immune escape to avelumab in a subset of MSS mCRC patients.
- These mutations may represent a specific patient subpopulation with unique selective pressures and treatment responses.
- Further trials of avelumab in MSS mCRC are warranted, particularly in this identified subpopulation.
