P2X7 receptors (P2X7R) are ATP-gated ion channels expressed in macrophages and dendritic cells (DC). Extracellular ATP, acting via P2X7R, is a highly efficacious stimulus for assembly of inflammasome signaling platforms that drive caspase-1 activation and secretion of biologically active interleukin-12 (IL-12). Inflammasome signaling is involved in an extraordinarily wide array of innate and adaptive immune responses to microbial pathogens or sterile host cell stresses, such as cancer. Recent studies have identified the P2X7R, and its ability to stimulate IL-12 secretion, as critical components of chemotherapy-induced anti-tumor cell immune responses. This response includes: 1) Primary chemotherapy induced-release of ATP from apoptotic tumor cells to activate P2X7R channels in DCs. 2) DC-mediated release of local IL-12 to tumor-reactive T cells that drives their polarization into anti-tumor-effector cells. 3) Lack of this immunogenic component of tumor cell killing in P2X7R-knockout mice which suffer from accelerated tumor growth during chemotherapy. These exciting findings raise several questions. How is P2X7R signaling in DCs initiated during their encounter with apoptotic cells at the tumor locus? What are the mechanisms that initiate and modulate release of ATP from the apoptotic tumor cells? We hypothesize that the efficacy of the P2X7R/IL-12 axis in chemotherapy-induced antitumor immune responses requires that these receptors act as sensors of not only ATP, but other extracellular metabolites, such as NAD and thiol reductants, which accumulate within apoptotic tumor microenvironments via the activation of metabolite-permeable channels. We will use in vitro and in vivo experimental models of murine DC function to test this hypothesis by addressing two major aims: 1) Define the purinergic signaling factors that regulate efficacy of the P2X7R/IL-12 signaling axis in DCs within the tumor microenvironment. These studies will characterize the roles of: a) P2X7R splice variants with distinctive channel gating and agonist selectivities;b) ART2 family ecto-ADP-ribosyltransferases that extracellular NAD to covalently modify and activate P2X7R;c) coactivated G protein-coupled P2Y2 nucleotide receptors or A2b adenosine receptors that may potentiate or attenuate P2X7R signaling and the anti-tumor immunogenic response. 2) Define the mechanisms underlying ATP/NAD release from apoptotic tumor cells and DCs which interact in the anti-tumor immunogenic axis. These studies will: a) quantify extracellular levels of ATP, ATP metabolites, and NAD within apoptotic tumor cell / DC co-cultures;b) characterize the roles of pannexin-1 channels and volume-regulated anion channels as conduits for ATP/NAD release from apoptotic thymocytes, apoptotic tumor cells, and DCs;c) define how ATP-permeable channel activities in apoptotic cells modulate the efficacy of P2X7R/IL-12 signaling within the anti-tumor immunogenic axis. These studies will provide mechanistic insights that may facilitate development of adjuvant purinergic receptor-based therapies which increase the efficacy of immunogenic anti-tumor responses to primary cancer chemotherapeutic agents.

Public Health Relevance

This study proposes to define the cellular signaling mechanisms by which the P2X7 purinergic receptor of inflammatory leukocytes can sense extracellular ATP and other metabolites released from dying tumor cells. Recent studies have identified the P2X7R as an important component of chemotherapy-induced anti-tumor cell immune responses. These studies will provide mechanistic insights that may facilitate development of adjuvant purinergic receptor-based therapies which increase the efficacy of immunogenic anti-tumor responses to primary cancer chemotherapeutic agents.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM036387-24
Application #
8446509
Study Section
Innate Immunity and Inflammation Study Section (III)
Program Officer
Nie, Zhongzhen
Project Start
1986-12-01
Project End
2015-02-28
Budget Start
2013-03-01
Budget End
2014-02-28
Support Year
24
Fiscal Year
2013
Total Cost
$366,404
Indirect Cost
$133,026
Name
Case Western Reserve University
Department
Physiology
Type
Schools of Medicine
DUNS #
077758407
City
Cleveland
State
OH
Country
United States
Zip Code
44106
Boyd-Tressler, Andrea; Penuela, Silvia; Laird, Dale W et al. (2014) Chemotherapeutic drugs induce ATP release via caspase-gated pannexin-1 channels and a caspase/pannexin-1-independent mechanism. J Biol Chem 289:27246-63
Dubyak, George R (2013) Dueling nucleosides: cross-regulation of extracellular adenosine by guanosine. Focus on "Extracellular guanosine regulates extracellular adenosine levels". Am J Physiol Cell Physiol 304:C403-5
Antonopoulos, Christina; El Sanadi, Caroline; Kaiser, William J et al. (2013) Proapoptotic chemotherapeutic drugs induce noncanonical processing and release of IL-1* via caspase-8 in dendritic cells. J Immunol 191:4789-803
Dubyak, George R (2011) Charge of the mito brigade. Focus on "Changes in mitochondrial surface charge mediate recruitment of signaling molecules during apoptosis". Am J Physiol Cell Physiol 300:C11-3
Villa-Bellosta, Ricardo; Wang, Xiaonan; Millan, Jose Luis et al. (2011) Extracellular pyrophosphate metabolism and calcification in vascular smooth muscle. Am J Physiol Heart Circ Physiol 301:H61-8
Qu, Yan; Misaghi, Shahram; Newton, Kim et al. (2011) Pannexin-1 is required for ATP release during apoptosis but not for inflammasome activation. J Immunol 186:6553-61
Trueblood, Katherine E; Mohr, Susanne; Dubyak, George R (2011) Purinergic regulation of high-glucose-induced caspase-1 activation in the rat retinal Muller cell line rMC-1. Am J Physiol Cell Physiol 301:C1213-23
Blum, Andrew E; Walsh, B Corbett; Dubyak, George R (2010) Extracellular osmolarity modulates G protein-coupled receptor-dependent ATP release from 1321N1 astrocytoma cells. Am J Physiol Cell Physiol 298:C386-96
Ramachandra, Lakshmi; Qu, Yan; Wang, Ying et al. (2010) Mycobacterium tuberculosis synergizes with ATP to induce release of microvesicles and exosomes containing major histocompatibility complex class II molecules capable of antigen presentation. Infect Immun 78:5116-25
Prosdocimo, Domenick A; Wyler, Steven C; Romani, Andrea M et al. (2010) Regulation of vascular smooth muscle cell calcification by extracellular pyrophosphate homeostasis: synergistic modulation by cyclic AMP and hyperphosphatemia. Am J Physiol Cell Physiol 298:C702-13

Showing the most recent 10 out of 76 publications