5-A-RU-PABC-Val-Cit-Fmoc is the precursor of 5-A-RU. 5-A-RU is the precursor of bacterial riboflavin, a mucosal-associated activator of invariant T (MAIT) cells. 5-A-RU forms potent MAIT-activating antigens through non-enzymatic reactions with small molecules from other metabolic pathways, such as glyoxal and methylglyoxal.

Physicochemical Properties

Molecular Formula	C43H53N9O13
Molecular Weight	903.93
Exact Mass	903.376
CAS #	2677841-58-4
PubChem CID	154573763
Appearance	White to off-white solid powder
LogP	0.8
Hydrogen Bond Donor Count	13
Hydrogen Bond Acceptor Count	14
Rotatable Bond Count	23
Heavy Atom Count	65
Complexity	1690
Defined Atom Stereocenter Count	5
SMILES	C(C1C2=CC=CC=C2C2=CC=CC=C12)OC(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCCNC(=O)N)C(=O)NC1C=CC(=CC=1)COC(=O)NC1C(NC(NC=1NC[C@H](O)[C@H](O)[C@H](O)CO)=O)=O
InChi Key	HGSKBBAITCBDNC-FAEFUHMUSA-N
InChi Code	InChI=1S/C43H53N9O13/c1-22(2)33(49-42(62)65-21-29-27-10-5-3-8-25(27)26-9-4-6-11-28(26)29)38(58)48-30(12-7-17-45-40(44)60)37(57)47-24-15-13-23(14-16-24)20-64-43(63)50-34-36(51-41(61)52-39(34)59)46-18-31(54)35(56)32(55)19-53/h3-6,8-11,13-16,22,29-33,35,53-56H,7,12,17-21H2,1-2H3,(H,47,57)(H,48,58)(H,49,62)(H,50,63)(H3,44,45,60)(H3,46,51,52,59,61)/t30-,31-,32+,33-,35-/m0/s1
Chemical Name	9H-fluoren-9-ylmethyl N-[(2S)-1-[[(2S)-5-(carbamoylamino)-1-[4-[[2,4-dioxo-6-[[(2S,3S,4R)-2,3,4,5-tetrahydroxypentyl]amino]-1H-pyrimidin-5-yl]carbamoyloxymethyl]anilino]-1-oxopentan-2-yl]amino]-3-methyl-1-oxobutan-2-yl]carbamate
HS Tariff Code	2934.99.9001
Storage	Powder-20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month Note: Please store this product in a sealed and protected environment, avoid exposure to moisture.
Shipping Condition	Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Biological Activity

ln Vitro

When added to MAIT cell cultures, 5-A-RU-PABC-Val-Cit-Fmoc (Compound 10) causes larger levels of mucosal-associated invariant T (MAIT) cell activation than 5-A-RU alone [1].

References

[1]. The Chemical Synthesis, Stability, and Activity of MAIT Cell Prodrug Agonists That Access MR1 in Recycling Endosomes. ACS Chem Biol. 2020 Feb 21;15(2):437-445. [2]. T-cell activation by transitory neo-antigens derived from distinct microbial pathways. Nature. 2014 May 15;509(7500):361-5. [3]. Recognition of Vitamin B Precursors and Byproducts by Mucosal Associated Invariant T Cells. J Biol Chem. 2015 Dec 18;290(51):30204-11. [4]. In Vitro and In Vivo Analysis of the Gram-Negative Bacteria-Derived Riboflavin PrecursorDerivatives Activating Mouse MAIT Cells. J Immunol. 2015 May 15;194(10):4641-9.

Solubility Data

Solubility (In Vitro)

DMSO: 150 mg/mL (165.94 mM)

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 3.75 mg/mL (4.15 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 37.5 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL. Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution. Solubility in Formulation 2: 3.75 mg/mL (4.15 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 37.5 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly. Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution. Solubility in Formulation 3: ≥ 3.75 mg/mL (4.15 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 37.5 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	1.1063 mL	5.5314 mL	11.0628 mL
	5 mM	0.2213 mL	1.1063 mL	2.2126 mL
	10 mM	0.1106 mL	0.5531 mL	1.1063 mL

*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.