Abstract 3034: Purine nucleoside phosphorylase regulates metabolic and immune checkpoints
Authors: Abt Evan R. , Rashid Khalid , Le Thuc M. , Lee Hailey R. , Creech Amanda L. , Wu Ting-Ting , Mehrling Thomas , Bantia Shanta , Radu Caius G. 
Published: June 2022

Abstract
Purine nucleoside phosphorylase (PNP) functions as a central regulator of nucleotide metabolism by catalyzing the degradation of purine nucleosides guanosine and deoxyguanosine. PNP inactivation has been paradoxically associated with both immunodeficiency and autoimmunity in humans but the genetic and molecular determinants of these divergent outcomes are poorly defined and whether they reflect cell type specific mechanisms is currently unknown. Moreover, the mechanisms underlying the clinical efficacy of PNP inhibitors in a subset of patients with hematological malignancies are yet to be determined, thereby severely limiting the potential of this promising new class of metabolic targeted therapies. To address these gaps in knowledge we performed a systematic evaluation of the metabolic, immunological and cell-context phenotypic effects of targeting PNP using a clinical-stage inhibitor with picomolar affinity and excellent oral bioavailability. Here we show that across cancer types PNP inhibition is synthetically lethal with sterile alpha motif and HD domain-containing protein 1 (SAMHD1) deficiency by triggering massive dGTP pool expansion, and impairing DNA replication. These cytotoxic effects require the expression of deoxycytidine kinase (dCK) which mediates the intracellular trapping of deoxyguanosine and expansion of dGTP pools, and are opposed by environmental deoxycytidine produced by bone marrow stromal cells in a SAMHD1 and nucleoside transporter dependent manner. This resistance mechanism to PNP inhibition can be counteracted by the deoxycytidine catabolizing enzyme cytidine deaminase (CDA).
We further show that in addition to, and independent of its dCK and CDA-dependent cytotoxic effects in SAMHD1 deficient cells, PNP inhibition elevates serum inflammatory cytokines and cytokine transcript levels in secondary lymphoid organs. Using scRNAseq we identify CD19+ B-cells and CSF1R+ macrophages, which express the pattern recognition receptor TLR7, as the drivers of these alterations. TLR7 activation in these cell types is promoted by guanosine accumulation which functions as a ligand for TLR7 alongside uridine-containing ssRNA. Importantly, TLR7 stimulation achieved by PNP inhibition initiates transcriptional alterations that are quantitatively and qualitatively unique from synthetic TLR7 agonists, such as the production of type I interferons. We additionally demonstrate that PNP inactivation stimulates germinal center responses in secondary lymphoid organs via the expansion of activated B-cells and T follicular helper cells. In summary, we demonstrate that by modulating multiple metabolic and immune checkpoints in a multigenic, cell type and environment dependent manner, pharmacological PNP inhibition elicits specific outcomes which can be leveraged for the development of novel therapeutic applications.
Citation Format: Evan R. Abt, Khalid Rashid, Thuc M. Le, Hailey R. Lee, Amanda L. Creech, Ting-Ting Wu, Thomas Mehrling, Shanta Bantia, Caius G. Radu. Purine nucleoside phosphorylase regulates metabolic and immune checkpoints [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3034.