Generation of effector Vγ9Vδ2 T cells and evaluation of their response to phosphoantigen-loaded cells

Hsiao, Chia‐Hung Christine; Wiemer, Andrew J.

doi:10.1016/j.xpro.2022.101422

Cited by 3 publications

(1 citation statement)

References 25 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The interferon-γ release was quantified using the previously reported protocol. 41 In T cell media, K562 cells were cultured with a density below 1 × 10 6 . The cells were split 1 day prior to the experiment.…”

Section: Methodsmentioning

confidence: 99%

Diester Prodrugs of a Phosphonate Butyrophilin Ligand Display Improved Cell Potency, Plasma Stability, and Payload Internalization

Singh,

Pawge,

Rani

et al. 2023

J. Med. Chem.

Self Cite

View full text Add to dashboard Cite

Activation of Vγ9Vδ2 T cells with butyrophilin 3A1 (BTN3A1) agonists such as ( E )-4-hydroxy-3-methyl-but-2-enyl diphosphate (HMBPP) has the potential to boost the immune response. Because HMBPP is highly charged and metabolically unstable, prodrugs may be needed to overcome these liabilities, but the prodrugs themselves may be limited by slow payload release or low plasma stability. To identify effective prodrug forms of a phosphonate agonist of BTN3A1, we have prepared a set of diesters bearing one aryl and one acyloxymethyl group. The compounds were evaluated for their ability to stimulate Vγ9Vδ2 T cell proliferation, increase production of interferon γ, resist plasma metabolism, and internalize into leukemia cells. These bioassays have revealed that varied aryl and acyloxymethyl groups can decouple plasma and cellular metabolism and have a significant impact on bioactivity (>200-fold range) and stability (>10 fold range), including some with subnanomolar potency. Our findings increase the understanding of the structure–activity relationships of mixed aryl/acyloxymethyl phosphonate prodrugs.

show abstract

Section: Methodsmentioning

confidence: 99%

Diester Prodrugs of a Phosphonate Butyrophilin Ligand Display Improved Cell Potency, Plasma Stability, and Payload Internalization

Singh,

Pawge,

Rani

et al. 2023

J. Med. Chem.

Self Cite

View full text Add to dashboard Cite

show abstract

Enhanced Plasma Stability and Potency of Aryl/Acyloxy Prodrugs of a BTN3A1 Ligand

Singh,

Pawge,

Rani

et al. 2024

ACS Med. Chem. Lett.

View full text Add to dashboard Cite

Mutations to the BTN2A1 Linker Region Impact Its Homodimerization and Its Cytoplasmic Interaction with Phospho-Antigen–Bound BTN3A1

Nguyen

Jin

Howell

et al. 2023

The Journal of Immunology

View full text Add to dashboard Cite

Intracellular binding of small-molecule phospho-Ags to the HMBPP receptor complex in infected cells leads to extracellular detection by T cells expressing the Vγ9Vδ2 TCR, a noncanonical method of Ag detection. The butyrophilin proteins BTN2A1 and BTN3A1 are part of the complex; however, their precise roles are unclear. We suspected that BTN2A1 and BTN3A1 form a tetrameric (dimer of dimers) structure, and we wanted to probe the importance of the BTN2A1 homodimer. We analyzed mutations to human BTN2A1, using internal domain or full-length BTN2A1 constructs, expressed in Escherichia coli or human K562 cells, that might disrupt its structure and/or function. Although BTN2A1 is a disulfide-linked homodimer, mutation of cysteine residues C247 and C265 did not affect the ability to stimulate T cell IFN-γ production by ELISA. Two mutations of the juxtamembrane region (at EKE282) failed to impact BTN2A1 function. In contrast, single point mutations (L318G and L325G) near the BTN2A1 B30.2 domain blocked phospho-Ag response. Size exclusion chromatography and nuclear magnetic resonance (NMR) experiments showed that the isolated BTN2A1 B30.2 domain is a homodimer, even in the absence of its extracellular and transmembrane region. [31P]-NMR experiments confirmed that HMBPP binds to BTN3A1 but not BTN2A1, and binding abrogates signals from both phosphorus atoms. Furthermore, the BTN2A1 L325G mutation but not the L318G mutation prevents both homodimerization of BTN2A1 internal domain constructs in size exclusion chromatography (and NMR) experiments and their binding to HMBPP-bound BTN3A1 in isothermal titration calorimetry experiments. Together, these findings support the importance of homodimerization within the BTN2A1 internal domain for phospho-Ag detection.

show abstract

Generation of effector Vγ9Vδ2 T cells and evaluation of their response to phosphoantigen-loaded cells

Cited by 3 publications

References 25 publications

Diester Prodrugs of a Phosphonate Butyrophilin Ligand Display Improved Cell Potency, Plasma Stability, and Payload Internalization

Diester Prodrugs of a Phosphonate Butyrophilin Ligand Display Improved Cell Potency, Plasma Stability, and Payload Internalization

Enhanced Plasma Stability and Potency of Aryl/Acyloxy Prodrugs of a BTN3A1 Ligand

Mutations to the BTN2A1 Linker Region Impact Its Homodimerization and Its Cytoplasmic Interaction with Phospho-Antigen–Bound BTN3A1

Contact Info

Product

Resources

About