Summary Intrahepatic cholangiocarcinoma (iCCA), the second most common primary liver cancer after hepatocellular carcinoma (HCC), is an aggressive malignancy with dismal overall prognosis. While frontline combination of chemotherapy with immune checkpoint inhibition (ICI) has been a major recent advance in therapy for patients with unresectable iCCA, there remains an unmet critical need to improve the current median progression free survival of about 8 months. With data supporting the need for a pre-existing immune response in the tumor for ICI response, here we propose to add high-dose conformal external beam radiotherapy (EBRT) followed by intra-tumor injection of autologous dendritic cells (DC) to dual PD-L1 (atezolizumab)/TIGIT (tiragolumab) blockade to further enhance the immune stimulatory effect. Radiation can induce inflammatory tumor cell death that can be favorable for tumor neoantigen presentation. Injection of autologous DC after EBRT would be a novel method of boosting in vivo tumor antigen uptake and presentation to expand tumor-reactive cytotoxic T cells. We have treated subjects with unresectable liver tumors (HCC and iCCA) in a pilot study with this EBRT and DC approach with promising response and acceptable toxicity (no grade ≥ 3 toxicity). Three of the 8 subjects had partial response, including an iCCA patient with ongoing response at 48 months. Both emergence of new T cell receptor (TCR) clones and expansion of existing TCR clones, including clones with tumor reactive and cytotoxic profile, have been observed, suggesting this combination could enhance tumor reactive cytotoxic T cell response. However, many of the TCR clones also have early exhaustion signal with upregulation of multiple checkpoint receptors including PD-L1 and TIGIT. Thus, combining DC injection with atezolizumab and tiragolumab may help further enhance the cytotoxic functions of these TCR clones. We hypothesize that combining EBRT followed by intratumor DC injections with atezolizumab and tiragolumab can improve the PFS for patients with unresectable iCCA and that the effect is mediated by systemic expansion of a tumor reactive T cell repertoire. We will test the hypothesis through 2 aims. 1) Assess the clinical efficacy of this combination therapy in a phase II study with a safety run-in phase. PFS will be the primary endpoint. 2) identify the effect of this novel combination immunotherapy on tumor reactive T cell repertoire. We will use scRNAseq and TCRseq to identify TCR clonal expansion and transcriptome profile of the TCR clones in the blood and tumor, with a focus on tumor reactive TCR clones. We will also use scRNAseq and flow to profile the changes of other immune cells in the tumor and blood. Finally, we will use imaging cytometry to examine the tumor and immune spatial relationship in the tumor. Our study will not only identify the clinical activities of this novel combination therapy but also use state-of-the-art technology to improve our understanding on the mechanism of action and potential resistance to this immunotherapy.

Narrative Patients with unresectable, intrahepatic cholangiocarcinoma have poor overall survival. We propose to examine the clinical activities and immune effect of a novel combination therapy using radiation followed by intratumor injection of autologous dendritic cells followed by systemic infusion of checkpoint inhibitors to PD-L1 and TIGIT.