Advancing Education, Research, and Quality of Care for the Head and Neck oncology patient.
Background: Recent advances in immuno-oncology have revolutionized cancer treatment; however, only a subset of patients achieve deep clinical responses. Immune evasion mechanisms that underlie resistance to these treatments remain poorly understood. Despite intensive research, the impact of regulatory T cells (Treg) as a predictor of immunotherapy response and survival in head and neck squamous cell carcinoma (HNSCC) remains elusive. Here, we investigate Treg transcriptional and spatial dynamics in HNSCC patients treated with neoadjuvant nivolumab (anti-PD-1, Nivo) alone or Nivo plus ipilimumab (anti-CTLA-4, Ipi) or relatlimab (anti-LAG-3, Rela) to uncover mechanisms by which immune checkpoint inhibitor (ICI) therapies may influence Treg and promote favorable clinical response. Clinical trial information: NCT04080804.
Methods: Pre- and post-treatment tumor samples from HNSCC patients (n=42) were processed for single-cell RNA-sequencing and TCR sequencing (scRNAseq), with sequencing successfully completed for 39 patients. Specific Treg marker genes and gene signatures were used to identify transcriptionally and clonally distinct Treg subpopulations. FFPE tissue sections were analyzed used multispectral imaging to assess changes in Treg infiltration and composition within the tumor and stroma.
Results: Increased CD3+ CD8- FOXP3+ Treg density in the stroma following Nivo+Ipi therapy was associated with a better pathologic response (p=0.018). In contrast, a higher density of Treg after Nivo+Rela regimen tended to correlate with a poorer pathologic response (p=0.13). There were no significant shifts in Treg density in the tumor with either treatment. Increased immune checkpoint receptor (ICR) expression by Treg in the tumor associated with major tumor rejection. Seven different transcriptomic clusters of tumor-infiltrating Treg were identified based on scRNAseq. A higher baseline presence of activated (p=0.03) and cycling Treg (p=0.01) correlated with major pathologic response to Nivo+Rela therapy. Moreover, activated Treg were modulated by Nivo+Rela and reprogrammed to a more naïve or memory-like phenotype in major responders.
Conclusion: Tumor infiltrating Treg influence clinical outcomes of neoadjuvant ICI therapy combinations in distinct ways. The presence of ICR-expressing Treg with an activated phenotype positively correlates with tumor rejection in response to Nivo+Rela treatment. These findings suggest a possible biologic basis for precision selection of ICI treatments.
Acknowledgements: This research was funded by BMS CA223049 (RLF), P50 CA097190 (RLF), R01 CA206517 (RLF), R01 DE031947 (RLF, LV, JHW), R01CA282074 (RLF and JHW) and DFG research fellowship #522169450 (MB). This research was partly supported by the University of Pittsburgh Center for Research Computing through the resources provided by using the HTC cluster, supported by NIH award number S10OD028483. This research utilized the UPMC Hillman Cancer Center Flow Cytometry Core Facility, partially supported by award P30 CA047904 (RLF).
Ethics Approval: This study was approved by the University of Pittsburgh’s Institutional Review Board; approval number HCC 18-139/CA224-056