Co-therapy with S-adenosylmethionine and nicotinamide riboside improves t-cell survival and function in Arts Syndrome (PRPS1 deficiency)

Abstract

Arts syndrome or phosphoribosyl-pyrophosphate-synthetase-1 (PRPS1) deficiency is caused by loss-of-function mutations in the PRPS1 gene (Xq22.3). PRPS1 is an initial and essential step for the synthesis of the nucleotides of purines, pyrimidines, and nicotinamide. Classically, affected males present with sensorineural hearing loss, optic atrophy, muscular hypotonia, developmental impairment, and recurrent severe respiratory infections early in life. Treatment of a 3-year old boy with S-adenosylmethionine (SAM) replenished erythrocyte purine nucleotides of adenosine and guanosine, while SAM and nicotinamide riboside co-therapy further improved his clinical phenotype as well as T-cell survival and function.

Keywords: 5-phosphoribosyl-1- pyrophosphate, PRPP; Adenosine triphosphate, ATP; Guanosine triphosphate, GTP; NAD phosphate, NADP; Nicotinamide riboside, NR; PRPP synthase, PRPPS; S-adenosylmethionine, SAM; nicotinamide adenine dinucleotide, NAD; phosphoribosyl-pyrophosphate-synthetase-1, PRPS1.

Fig. 1

Three-dimensional structure of the PRPPS and environment of the p.Arg84Trp site. (A) The PRPPS hexamer as assembled from PDB entry 2HCR, which contains AMP (sticks in magenta) and a Cd2+ ion (magenta sphere) in the ATP binding site, and sulfate ions in the ribulose-5-phosphate (R5P, black sticks) binding site and allosteric sites I and II (space fill models in black and dark grey, respectively). Each monomer is coloured differently, but similar shades were used for those forming the more tightly interacting homodimers. A space-fill model of the Arg84 side chain in red indicates the variant site. For clarity, the ligands and variant site are labelled in one of the subunits only. (B) Close-up stereo views of the p.Arg84Trp substitution site. The side chain conformation causing the least steric clashes is chosen for the introduced Trp84 shown as stick model with carbon atoms in magenta. The replaced Arg84 side chain is shown with thinner sticks and carbon atom color corresponding to that of the respective subunit. Details of the hydrogen bonding interactions of Arg84 and of the sulfate ion bound at allosteric site I (labelled I) are shown for one of the subunits. Residues hydrogen bonding to Arg84 or sterically clashing with the variant-introduced Trp84 side chain are hereby depicted with carbon atoms in yellow, whereas sulfate ion-binding residues are shown with carbon atoms in white. The latter originate from three different subunits, as indicated by one, two or no apostrophes in the residue label. Hydrogen bonds are depicted as dashed lines. The nearby allosteric site II is labelled II and contains a sulfate ion shown as space fill model in dark grey.

Fig. 2

Nucleotide levels in erythrocytes in the patient under different treatment conditions and in healthy controls. Values of controls are given as means with standard deviations [24].

Fig. 3

The patient’s T cells have diminished cytokine responses and survival, which are partially or fully rescued by treatment with SAM + NR. CD8 T cells from three healthy controls (two adult and one adolescent (16 y.o. designated with #)) and from the patient after 24 hours of treatment washout (- tx) or during treatment (+ tx) were activated with CD3 + CD46 (CD46 co-stimulation is important for optimal human effector CD8 T cell responses). (A) IFN-ɣ and TNF-α secretion was assessed in the cell supernatants at ~40 hours post-activation (1° response) and after rest and re-stimulation overnight (2° response). At 6 days post-activation (B) and 5 days after rest (C) the number of live cells was enumerated. Proliferation was assessed by intracellular Ki67 expression (D) and the percentage of dead cells was calculated (E) after the 1° and 2° response using flow cytometric analysis.

Fig. 4

The patient’s CD4 T cells have diminished cytokine responses and survival, which are partially rescued by treatment with SAM + NR. CD4 T cells from three healthy controls (two adult and one adolescent (16 y.o. designated with #)) and from the patient after 24 hours of treatment washout (- tx) or during treatment (+ tx) were activated with CD3 + CD46, a necessary co-stimulator for human Th1 responses (CD46 deficient patients do not mount Th1 responses). (A) IFN-g and TNF-a secretion was assessed in the cell supernatants after activation, rest and overnight re-stimulation. (B) At 6 days post-activation the number of live cells was enumerated. The percentage of dead cells was calculated after the (C) primary stimulation and (D) secondary stimulation (overnight re-stimulation) using flow cytometric analysis.

Fig. 5

The patient's CD4 T cells have diminished IFN-γ cytokine responses, which are rescued by combined treatment with SAM + NR, but not with SAM alone. CD4 T cells from four healthy controls (naïve CD4 T cells from two adults and total CD4 T cells from a 16 and 17 year old) and the patient during SAM treatment only or SAM + NR treatment and activated with CD3 + CD46, a necessary co-stimulator for human Th1 responses (CD46 deficient patients do not mount Th1 responses). (A) IFN-γ secretion was assessed in the supernatants at ~40 h post-activation. (B) CD4 T cells from a healthy control and the patient during SAM + NR treatment were activated with CD3 + CD46 in the presence of vehicle control or 25 nM or 100 nM of SAM and IFN-γ secretion was assessed at ~40 h post-activation.