Nucleotide metabolism in cancer cells fuels a UDP-driven macrophage cross-talk promoting immunosuppression and immunotherapy resistance [RNAseq_Panc02_sgNT_sgCda_tumor]

Purpose: Here we use Here, we used RNA sequencing to identify trascriptictomic profile of sgNT and sgCda PancO2 tumor bearing-mice. Methods: 4x10e6 sgNT and sgCda Panc02 cells were injected subcutaneously (s.c.) in the right flank of the C57BL/6J mouse in a final suspension of 200 μL PBS. Tumor volumes were measured at least three times per week. At day 16 after injection, tumor-bearing mice were sacrificed by cervical dislocation and RNA was isolated from tumor tissue using TRIzol (Life Technologies). Starting from total RNA, poly-adenylated fragments were isolated, reverse transcribed and converted into indexed sequencing libraries using the KAPA stranded mRNA-seq kit (Sopachem, Eke, Belgium). The first 50 bases of these libraries were sequenced on a HiSeq 4000 system (Illumina, San Diego, CA). These raw reads were mapped after removal of the sequencing adapters to the reference transcriptome and genome (GRCm38/mm10) using the Bowtie 2.0 and TopHat 2.0 pipeline (Langmead and Salzberg, 2012).