GDNPs reprogram macrophages to regulate arginase-1 release for ameliorating T cell exhaustion in tumor microenvironment

Lines of evidence indicate that, immune checkpoints (ICs) inhibitors enhance T cell immune response to exert anti-tumor effects. However, T cell exhaustion is still a major obstacle to antitumor immunotherapy in colorectal cancer patients. Our previous studies showed that ginseng-derived nanoparticles (GDNPs) inhibited the growth of various tumors by reprograming tumor-associated macrophages (TAMs) and downregulated the ICs expression on T cells in tumor microenvironment (TME), but the underlying effector mechanisms remained unclear. In this study, we demonstrated that GDNPs reprogramed TAMs via reducing arginase-1 (ARG1) production. Moreover, normalized arginine metabolism ameliorate T cell exhaustion through mTOR-T-bet axis, resulting in reduced ICs expression and enhanced CD8+ T cells expansion. Together, these results provide new insights in understanding the mechanisms involved in reversing T cell exhaustion, which may facilitate the development of new therapeutic strategy for cancer treatment. Overall design: Single cell RNA seqencing data of six samples from six colon cancer bearing C57 mice treated with ginseng-derived nanoparticles. Each samples contain one sample from solid tumor of each tumor bearing mouse. Mouse tumor tissues were scissored into 5 mm pieces and incubation with 100 µg/mL Liberase and 0.2 mg/mL DNase ? in PBS for 45 min at 37°C. After digestion, 70 µm nylon net was centrifuged at 300 g for 5 min. Subsequently, the supernatants were discarded, and the cell pellets were suspended in 1 mL PBS. To remove red blood cells, 2 mL of GEXSCOPETM red blood cell lysis buffer (Singleron) was added, and cells were incubated at 25 °C for 10 min. The solution was then centrifuged at 500 × g for 5 min and resuspended in PBS.Single-cell suspensions (1×105 cells/mL) with PBS were loaded into microfluidic devices using the Singleron Matrix®Single Cell Processing System (Singleron). Subsequently, the scRNA-seq libraries were constructed according to the protocol of the GEXSCOPE® Single Cell RNA Library Kits (Singleron). Individual libraries were diluted to 4 nM and pooled for sequencing. At last, pools were sequenced on Illumina HiSeq X with 150 bp paired end reads.