Vγ9Vδ2 T cell activation by strongly agonistic nucleotidic phosphoantigens

Abstract

Human Vγ9Vδ2 T cells can sense through their TCR tumor cells producing the weak endogenous phosphorylated antigen isopentenyl pyrophosphate (IPP), or bacterially infected cells producing the strong agonist hydroxyl dimethylallyl pyrophosphate (HDMAPP). The recognition of the phosphoantigen is dependent on its binding to the intracellular B30.2 domain of butyrophilin BTN3A1. Most studies have focused on pyrophosphate phosphoantigens. As triphosphate nucleotide derivatives are naturally co-produced with IPP and HDMAPP, we analyzed their specific properties using synthetic nucleotides derived from HDMAPP. The adenylated, thymidylated and uridylated triphosphate derivatives were found to activate directly Vγ9Vδ2 cell lines as efficiently as HDMAPP in the absence of accessory cells. These antigens were inherently resistant to terminal phosphatases, but apyrase, when added during a direct stimulation of Vγ9Vδ2 cells, abrogated their stimulating activity, indicating that their activity required transformation into strong pyrophosphate agonists by a nucleotide pyrophosphatase activity which is present in serum. Tumor cells can be sensitized with nucleotide phosphoantigens in the presence of apyrase to become stimulatory, showing that this can occur before their hydrolysis into pyrophosphates. Whereas tumors sensitized with HDMAPP rapidly lost their stimulatory activity, sensitization with nucleotide derivatives, in particular with the thymidine derivative, induced long-lasting stimulating ability. Using isothermal titration calorimetry, binding of some nucleotide derivatives to BTN3A1 intracellular domain was found to occur with an affinity similar to that of IPP, but much lower than that of HDMAPP. Thus, nucleotide phosphoantigens are precursors of pyrophosphate antigens which can deliver strong agonists intracellularly resulting in prolonged and strengthened activity.

Improving the Efficiency of Vγ9Vδ2 T-Cell Immunotherapy in Cancer

“…Activation of Vγ9Vδ2 T-cells with PAgs and N-BPs is the most commonly used strategy for in vitro research and both in vivo stimulation as well as application of adoptive cell therapy. The potency of the individual PAg molecule to elicit response from Vγ9Vδ2 T-cells differs ( 108 ) and is especially high for microbial (E)-4-hydroxy-3-methyl-butenyl pyrophosphate (HMBPP), certain synthetic compounds like bromohydrin pyrophosphate (BrHPP) ( 109 ) or nucleotides derived from HMBPP ( 110 ). However, so far only BrHPP and N-BPs have been used clinically.…”

“…Nano-technology based carriers for N-BP delivery ( 141 ) as well as lipophilic bisphosphonate ( 60 , 142 , 143 ) and synthetic nucleotide pyrophosphates ( 110 ) are additional pharmacotherapeutic strategies that may improve Vγ9Vδ2 T-cell immunotherapy in the future.…”

Aiming for the Sweet Spot: Glyco-Immune Checkpoints and γδ T Cells in Targeted Immunotherapy

“…To activate the Vγ9Vδ2 T-cells, the butyrophilin-mediated reactivity of the Vγ9Vδ2 TCR to phosphoantigens can be exploited, using chemical compounds to elevate or mimic the expression of phosphoantigens either on tumor cells or on antigen-presenting cells in the TME. Such compounds include aminobisphosphonates (for example pamidronate and zoledronate) or synthetic phosphoantigen analogs ( 197 , 198 ). The approach offers a useful tool for expansion, but is not necessarily a useful therapeutic approach, because the expanded Vγ9Vδ2 T cells are nearly monoclonal and are not specific for a desired antigen.…”

Towards Deciphering the Hidden Mechanisms That Contribute to the Antigenic Activation Process of Human Vγ9Vδ2 T Cells

“…These conformational changes could represent the first signals delivered to distinguish activating or non-activating molecules. In fact, the BTN3A1 B30.2 domain is able to bind additional negatively charged small molecules, like malonate, citrate, adenosine-diphosphate (ADP), and nucleotidic pAg ( 59 ). Exogenous pAg (e.g., HMBPP/HDMAPP) induce a chemical shift into the B30.2 domain that extended to the binding site.…”