Potentiating radiation-induced T1IFN and the anti-tumoral immune response with the ATM inhibitor AZD1390 in pancreatic cancer
Radiotherapy induces a Type I interferon (TIFIN)-mediated anti-tumoral immune response that we hypothesized could be potentiated by a first-in-class ATM inhibitor leading to enhanced innate immune signaling. T1IFN expression, and sensitization to immunotherapy in pancreatic cancer. We evaluated the effects of AZD1390 or a structurally related compound AZD0156 on innate immune signaling and found that both inhibitors enhanced radiation-induced  T1IFN expression via the POLIII/RIG-I/MAVS pathway. In immunocompetent syngeneic mouse models of pancreatic cancer, ATM inhibitor enhanced radiation-induced anti-tumoral immune responses and sensitized to anti-PD-L1, producing immunogenic memory and durable tumor control. Therapeutic responses were associated with increased CD8+ T cell frequency and effector function. Tumor control was dependent on CD8+ T cells as therapeutic efficacy was blunted in immunodeficient or CD8+ T cell-depleted mice. Adaptive immune responses to combination therapy provided systemic control of contralateral tumors outside of the radiation field. Taken together, we show that a clinical candidate ATM inhibitor enhances radiation-induced T1IFN leading to both innate and subsequent adaptive immune response and sensitization of otherwise resistant pancreatic cancer to immunotherapy. Overall design: 6-8 week old C57/Bl6 mice with subcutaneous mT4 tumors were treated with AZD1390 (20mg/kg) for 5 days starting on day 0, 1 dose of radiation (8Gy) on day 0, and 3 doses of anti-PD-L1 blocking antibody (100µg) once every three days starting on day -1. 6-8 tumors were harvested for each of the following treatment groups: IgG/control, AZD1390, RT+anti-PD-L1, and AZD1390+RT+anti-PD-L1. Tumors were harvested on day 5 and digested in a collagenase digest buffer, and filtered through 40µm mesh filter. Samples were depleted of dying cells twice using Dead Cell Removal Kit from Miltenyi. Cells were then seperated based on CD45 status, and mixed back in equal parts CD45+ and CD45- to enrich immune cells present in the samples. Samples were diluted to a final concentraiotn of 700-1000 cells/ul. Single cell sequencing was performed by the University of Michigan Advanced Genomics Core Research Facility. Library generation was performed on the 10x Genomica Chromium Controller following manufacturers protocol for the 3'v3.1 chemistry with NextGEM Chip G reagents. Library quality was assess by LabChip GXII HT and quantified by Qubit. Pooled libraries were subjected to 150 base pair, paired-end sequencing according to manufacturer protocol (Illumina NovaSeq 6000). Bcl2fastq2 Conversion Software was used to de-multiplex fastq files and CellRanger v7 Pipeline was used to align reads and generate count matrices.