Inhaled TLR9 Agonist Renders Lung Tumors Permissive to PD-1 Blockade Promoting Optimal CD4+ and CD8+ T cell Interplay

Currently approved inhibitors of the PD-1/PD-L1 pathway represent a major advance for the treatment of lung cancers, yet they are ineffective in a majority of patients due to lack of pre-existing T cell reactivity. Here we show that a TLR9 agonist delivered by inhalation is able to prime T cell responses against poorly immunogenic lung tumors and to complement the effects of PD-1 blockade. Treatment with inhaled TLR9 agonist causes profound remodeling in tumor-bearing lungs, leading to formation of tertiary lymphoid structures adjacent to the tumors, CD8+ T cell infiltration into the tumors, dendritic cell expansion and antibody production. Inhaled TLR9 agonist treatment increased the pool of functional PD-1lowT-bethigh effector CD8+ T in tumor-bearing lungs. We show by transcriptional profiling, that the effector CD8+ T cells generated by inhaled TLR9 agonist treatment are licensed by PD-1 blockade to become highly functional CTL, leading to a durable rejection of both lung tumors and tumor lesions outside the lungs. CD4+ T cells activated in response to inhaled TLR9 play a critical role in this process by controlling the proliferation, preventing exhaustion, and guiding the differentiation of optimally functional CTL. This study characterizes a strategy to apply localized TLR9 stimulation to a tumor type not accessible for direct injection, a strategy that may expand the therapeutic potential of PD-1 blockade in non-small cell lung cancer. the scope of this experiment was to profile the gene expression of effector CD8 T cells from tumor bearing lungs treated by different drug regimen. Gene expression was performed on purified CD44+CD8+ effector T cells from 4T1 tumor-bearing lungs treated with the indicated treatment regimen (group names reflect treatment). Saline-CTRL (n=5), SD-101 (n=6), SD-101+anti-PD-1 (n=5), SD-101+anti-PD-1+a-CD4 (n=4)