Cross-presentation by type 1 DCs (cDC1) is critical to induce and sustain antitumoral CD8 T cell responses to model antigens, in various tumor settings. However, the impact of cross-presenting cDC1 and the potential of DC-based therapies in tumors carrying varied levels of bona-fide neoantigens (neoAgs) remain unclear. Here we develop a hypermutated model of non-small cell lung cancer in female mice, encoding genuine MHC-I neoepitopes to study neoAgs-specific CD8 T cell responses in spontaneous settings and upon Flt3L + αCD40 (DC-therapy). We find that cDC1 are required to generate broad CD8 responses against a range of diverse neoAgs. DC-therapy promotes immunogenicity of weaker neoAgs and strongly inhibits the growth of high tumor-mutational burden (TMB) tumors. In contrast, low TMB tumors respond poorly to DC-therapy, generating mild CD8 T cell responses that are not sufficient to block progression. scRNA transcriptional analysis, immune profiling and functional assays unveil the changes induced by DC-therapy in lung tissues, which comprise accumulation of cDC1 with increased immunostimulatory properties and less exhausted effector CD8 T cells. We conclude that boosting cDC1 activity is critical to broaden the diversity of anti-tumoral CD8 T cell responses and to leverage neoAgs content for therapeutic advantage.

The efficacy of immune checkpoint blockade (ICB) in NSCLC depends on the tumor mutational burden (TMB). However, a fraction of patients with high TMB and predicted immunogenic neoantigens (neoAgs) do not respond. Here we show that in a model of highly mutated NSCLC, cross-presenting cDC1s are required to induce broad effector CD8+ T cell responses to both strong and weak endogenous neoAgs. Importantly, cDC1 amplification by Flt3L increases immunogenicity of MHC class-I neoepitopes and promotes tumor regression, whilst PD-L1 blockade is ineffective. cDC1 density correlates to CD8+ T cell scores and prognosis, particularly in hypermutated human NSCLC. Single-cell RNA sequencing reveals the molecular determinants of Flt3L-therapy including expansion of immunogenic lung cDC1 and proliferation of cytotoxic CD8+ T cells with reduced exhaustion. We conclude that boosting cDC1 activity is critical to leverage neoAgs content for therapeutic advantage in hypermutated lung tumors that do not respond to ICB.