Lenvatinib plus Anti-PD-1 Combination Therapy for Advanced Cancers: Defining Mechanisms of Resistance in an Inducible Transgenic Model of Thyroid Cancer.

Background: Combination therapy with lenvatinib plus programmed death-1 (PD-1) immune checkpoint blockades (ICB) is under investigation in many solid tumors, including thyroid cancer. Lenvatinib is known to reduce angiogenesis and may overturn the immune-suppressive effects of VEGF in the tumor microenvironment. Previous studies investigating the effects of VEGFR inhibition on the immune response were performed in rapidly growing tumor models where immune equilibrium is not established prior to treatment. We hypothesize that physiologically-relevant preclinical models are necessary to define mechanisms of resistance to immune-targeted combination therapies. Methods: We utilized the TPO-CreER/BrafV600E/wt/Trp53Δex2-10/Δex2-10 inducible transgenic model of advanced thyroid cancer to investigate lenvatinib treatment in the context of an anti-PD-1 ICB. Following tumor establishment, 3.5 months post-induction, mice were treated with high (10mg/kg) or low-dose (2mg/kg) lenvatinib, anti-PD-1, or combination lenvatinib with anti-PD-1. Tumor volume and lung metastases were assessed in each group. Immune infiltrate was characterized by flow cytometry and immunohistochemistry, and TCRβ sequencing was performed to further investigate the T cell response. Results: Both low and high-dose lenvatinib reduced tumor volume, while anti-PD-1 alone had no effect alone or in combination. Although both low and high-dose lenvatinib reduced vascular density, low-dose was superior in controlling tumor size. Lung metastases and survival were not improved with therapy, despite the effects of lenvatinib on primary tumor size. Low-dose lenvatinib treatment led to a subtle reduction in the dominant Ly6G+CD11b+ myeloid cell population and was associated with increased CD4+ T cell infiltrate and an enrichment in 4-1BB+ and granzyme B+ CD4+ T cells and FoxP3+ Tregs. Polyclonal T cell expansions were evident in the majority of mice, suggesting that a tumor-specific T cell response was generated. Conclusions: The effects of lenvatinib on the immune response were most pronounced in mice treated with low-dose lenvatinib, suggesting that dose should be considered in clinical application. While the immune-modulating potential of lenvatinib is encouraging, alterations in the immune milieu and T cell activation status were insufficient to sustain durable tumor regression, even with added anti-PD-1. Additional studies are necessary to develop more effective combination approaches in low-mutation burden tumors, such as thyroid cancer.