5-Amino-1MQ: Quinolinium Chemistry and NNMT Enzyme Inhibition

5-Amino-1MQ is built on quinoline, a bicyclic aromatic system of fused benzene and pyridine rings. Methylation of the pyridine-type nitrogen converts that atom into a quaternary ammonium centre carrying a fixed positive charge, while an amino group occupies carbon 5 on the carbocyclic ring. Formula C10H11N2+, monoisotopic mass 159.09222 daltons.

Because the nitrogen is quaternised rather than protonated, the charge cannot be titrated away. There is no basic pKa to cross and no neutral species to form, which is why the material is always isolated as a salt with a halide counter-ion. PubChem catalogues the iodide as CID 66522933 (C10H11IN2, 286.11 g/mol, CAS 42464-96-0) and the chloride as CID 176507677 (C10H11ClN2, 194.66 g/mol). The bare cation is CID 950107, InChIKey ZMJBCEIHNOWCMC-UHFFFAOYSA-O.

The computed descriptors are unusually spare for a molecule with measurable enzyme potency. PubChem reports XLogP 1.1, a topological polar surface area of 29.9 square angstroms, one hydrogen-bond donor, one acceptor and zero rotatable bonds. A rigid, flat, singly charged bicycle of that size has almost nowhere to hide conformational entropy, so essentially all of its affinity has to come from shape complementarity and electrostatics inside the pocket it occupies.

The shorthand name decomposes cleanly. 1MQ is 1-methylquinolinium, the scaffold; the 5-amino prefix names the substituent and its ring position. That parent scaffold is itself familiar in this field for a separate reason: 1-methylquinolinium is the product NNMT forms when it methylates quinoline, which is what makes quinoline a convenient surrogate substrate in enzyme assays.

5-Amino-1MQ acts on nicotinamide N-methyltransferase (EC 2.1.1.1), a 264-residue cytosolic protein of 29,574 daltons encoded by the NNMT gene and catalogued in UniProt as P40261. The enzyme transfers a methyl group from S-adenosyl-L-methionine to the ring nitrogen of nicotinamide, releasing 1-methylnicotinamide and S-adenosyl-L-homocysteine.

Expression is heavily tissue-weighted. UniProt records the protein as predominantly hepatic, with lower amounts in kidney, lung, skeletal muscle, placenta and heart, and no detection in brain or pancreas. Substrate scope is broader than the name implies: NNMT accepts pyridine and quinoline analogues as well as nicotinamide itself, which is the basis of the surrogate-substrate assays described further down.

Reported steady-state constants place the cofactor well ahead of the substrate in affinity. A 2021 mechanistic review quotes a Michaelis constant of 199 plus or minus 32 micromolar for nicotinamide against 8.5 plus or minus 0.8 micromolar for S-adenosyl-L-methionine, roughly a twentyfold separation, and describes a rapid-equilibrium ordered bi-bi sequence in which the cofactor binds first and products leave in reverse order. A 2017 real-time fluorescence study reached a different conclusion from global curve fitting across a substrate concentration array, favouring a random bireactant model in which either substrate can bind the free enzyme, with each then binding the complementary binary complex about twentyfold more tightly than the apoenzyme. Both descriptions agree on the practical consequence: the two sites are energetically coupled, so occupancy at one changes affinity at the other.

The reaction sits at a junction between two metabolite pools. Methylation removes nicotinamide from the pool available to the NAD+ salvage route and simultaneously consumes a methyl equivalent from S-adenosyl-L-methionine, and the 1-methylnicotinamide produced is exported rather than recycled. Reviews from 2026 describe this as a methyl sink, and it is the reason NNMT catalytic output is usually read out by measuring 1-methylnicotinamide rather than by measuring the enzyme itself.

5-Amino-1MQ occupies the nicotinamide substrate pocket rather than the cofactor site, according to the docking work published alongside the original potency series. Predicted binding orientations placed the quinolinium ring against substrate-site residues, and ligand-enzyme interaction scores across the analogue set correlated closely with the inhibition constants measured experimentally.

The structural frame for that pocket was set by crystallography. PDB entry 2IIP resolved human NNMT with S-adenosyl-L-homocysteine bound at 2.05 angstroms, and Peng and colleagues reported the ternary picture in Biochemistry in 2011, solving the enzyme with both S-adenosyl-L-homocysteine and nicotinamide at 2.7 angstroms (PDB 3ROD). Mutagenesis in that paper identified aspartate 197 and tyrosine 20 as the load-bearing active-site residues: substituting either dropped the catalytic efficiency term by two to three orders of magnitude, raising the Michaelis constants for both nicotinamide and the cofactor while lowering the turnover number only modestly. Molecular dynamics accounted for the long-range effect by which the D197A change perturbed cofactor binding from a distance.

Around those two residues, the mechanistic literature repeatedly names tyrosine 25, aspartate 85, tyrosine 86, asparagine 90, aspartate 142, valine 143, alanine 169, serine 201, tyrosine 204 and serine 213 as contact points, with cysteine 165 exploited separately as a handle for covalent chemistry. The adenine ring of the cofactor packs into a hydrophobic slot formed by tyrosine 86 and alanine 169 while hydrogen bonding to aspartate 142 and valine 143, and the ribose hydroxyls hydrogen bond to aspartate 85 and asparagine 90. The UniProt entry now lists nearly thirty deposited human NNMT structures, a substantial fraction of them inhibitor complexes.

Structure-activity work across the scaffold family explains why the 5-amino substitution matters. Screening N-methylated quinolinium, isoquinolinium, pyridinium and benzimidazolium or benzothiazolium analogues singled out quinoliniums as the productive scaffold, and small substituents such as methyl or amino were tolerated on the ring while bulkier groups lost affinity to steric clash. The primary amine at position 5 is close to the largest change the pocket will accept without penalty.

5-Amino-1MQ sits in the low micromolar band of NNMT inhibition, with a reported IC50 of 1.2 micromolar against recombinant human enzyme assayed at 50 micromolar S-adenosyl-L-methionine and 100 micromolar substrate. Two close relatives are roughly an order of magnitude weaker: unsubstituted 1-methylquinolinium at 12.1 micromolar and the reaction product 1-methylnicotinamide at 9.0 micromolar.

A second chemotype reaches far lower numbers by covering both sites at once. Bisubstrate inhibitors tether a nicotinamide-like fragment to an adenosine-like fragment through a linker that threads the methyl transfer tunnel. Policarpo and colleagues built NS1 around an alkyne linker chosen to mimic the linear 180-degree geometry of the transition state, synthesised it in fourteen steps and reported subnanomolar potency, tabulated at 500 picomolar in a later review. Iyamu and colleagues subsequently reported II559 and II802 with inhibition constants of 1.2 and 1.6 nanomolar and better than 5000-fold selectivity over related methyltransferases.

The trade-off is transport, not affinity. Bisubstrate compounds carry the charge and polarity of two nucleotide-like fragments, so biochemical potency in the nanomolar range routinely collapses in intact cells. II559 and II802 reached roughly 150 nanomolar in cells, and a separate 2021 study had to esterify both the amine and the carboxylate of the amino acid side chain to get a single-digit nanomolar bisubstrate inhibitor across a membrane at all, with the isopropyl ester giving the best combination of buffer stability and esterase-dependent release of the parent compound. Against that background the appeal of a 159-dalton monocation with an XLogP near 1 is straightforwardly physicochemical.

Recent chemotypes have narrowed the gap from the other direction. A 2024 series modified the adenine N7 position to a 7-deazaadenosine and tuned the linker, reaching an IC50 of 47.9 plus or minus 0.6 nanomolar with clean selectivity across a human methyltransferase panel. A 2025 piperazine-based series combining quinolone and pyridine pharmacophores reported nanomolar potency for compound 7b with inhibition competitive against both the cofactor and the quinoline substrate. And a 2026 report described non-SAM-mimetic bisubstrate inhibitors found by high-throughput screening, with the lead improving more than a thousandfold over the original hit to sub-micromolar cellular activity.

5-Amino-1MQ belongs to a methylquinolinium series that was profiled against structurally related methyltransferases and against the enzymes of the NAD+ salvage route. Counter-screening found no inhibition of those off-target enzymes, and permeability panels showed that analogues carrying a primary amine substituent crossed both artificial membranes and Caco-2 monolayers by passive and carrier-mediated routes.

In cultured adipocytes the same series lowered intracellular 1-methylnicotinamide, the direct catalytic product, while intracellular NAD+ and S-adenosyl-L-methionine rose. Measuring the product rather than a downstream phenotype is what makes that readout a target-engagement measure: 1-methylnicotinamide has no route back into the salvage pathway, so its concentration tracks flux through the enzyme.

The transport question deserves separating from the potency question. A permanent monocation does not diffuse across a bilayer the way a neutral drug-like molecule does, and organic cation transporters of the OCT and MATE families are the recognised handlers of small quaternary species; 1-methylnicotinamide itself is well established as an OCT2 and MATE substrate and is used as an endogenous probe for those carriers in transporter studies. The permeability data above report that both passive and active routes contribute for the aminated methylquinoliniums, without resolving which specific carriers are involved.

Cellular consequences of NNMT inhibition have been mapped most thoroughly in disease-model systems rather than with this compound in particular. A 2025 Nature paper traced NNMT activity in cancer-associated fibroblasts to hypomethylation of histone H3 lysine 27, which drove complement secretion and recruitment of myeloid-derived suppressor cells, with genetic loss of the enzyme restoring CD8+ T cell activation across syngeneic tumour models. A 2026 Cell Reports study using spatial transcriptomics on human kidney biopsy tissue linked NNMT-positive tubules to senescence and fibrosis signatures, and found that enzyme overexpression worsened epithelial-to-mesenchymal transition in tubular cells while selective inhibition was protective in organoids. Both illustrate the same methodological point: the phenotype is read out downstream, and the enzymological measurement stays separate.

5-Amino-1MQ potency is most often measured with a noncoupled fluorescence assay that swaps quinoline in for nicotinamide, because the methylated product 1-methylquinolinium fluoresces with an emission maximum near 405 nanometres while quinoline itself excites poorly beyond 320 nanometres. Reported detection extends to roughly 40 nanomolar product when the reaction buffer is dilute Tris.

The value of a direct readout is that it removes the coupling enzymes that most methyltransferase assays use to convert S-adenosyl-L-homocysteine into a detectable signal. Coupled formats report on a two-enzyme cascade, so any compound that touches the auxiliary enzymes registers as a false positive. A single-step fluorescence readout follows product appearance in real time, which is what allowed the 2017 study to fit initial-velocity data across a grid of substrate concentrations and argue about the reaction mechanism from the fit quality. The same format generates concentration-response curves quickly enough for library screening.

Alternative formats fill in what fluorescence cannot cover. Fluorescence polarisation competition assays report displacement from the active site rather than turnover. Liquid chromatography with tandem mass spectrometry quantifies 1-methylnicotinamide directly, which is the standard route for cellular and tissue samples; validated bioanalytical methods for that metabolite in human plasma and urine reach lower limits near 2 nanograms per millilitre against basal plasma concentrations of roughly 4 to 120 nanograms per millilitre.

The most detailed kinetic dissection to date came from surface biosensing. A 2025 Journal of Biological Chemistry paper reported a biosensor method that quantifies substrate binding kinetics to the enzyme-cofactor complex, the catalytic rate and the rate of product release from the enzyme-homocysteine complex in a single experiment, described by the authors as the first use of an enzyme surface biosensor to resolve catalysis at that level of detail. The same work established that a class of nicotinamide-analogue inhibitors is itself turned over by the enzyme, with the methylated product acting as the potent species, so on-target biotransformation converts a permeable precursor into a charged, high-affinity inhibitor inside the cell.

5-Amino-1MQ is supplied as a crystalline halide salt, most commonly the iodide (CAS 42464-96-0, formula mass 286.11), with the chloride form also catalogued. Research-grade material from chemical suppliers is specified at 98 percent or greater purity by high-performance liquid chromatography, and identity is confirmed by proton nuclear magnetic resonance and mass spectrometry.

Mass spectrometric identity is unusually simple for this compound. Because the analyte is already a cation, electrospray in positive mode detects the intact species without adduct formation, at a monoisotopic m/z of 159.0922 for C10H11N2+. The counter-ion is invisible to the detector, so the same spectrum is obtained from the iodide and the chloride, and salt form has to be established separately by elemental analysis or ion chromatography.

Storage practice follows from the chemistry rather than from any published stability study specific to this molecule. A quaternary ammonium halide is hygroscopic, so desiccated storage keeps the assayed mass honest; iodide salts are additionally photolabile, since iodide oxidises to iodine on light exposure and colours the solid, which is why amber glass and low-temperature storage are the default for the iodide form. Aromatic primary amines are oxidation-prone in solution, so freshly prepared aqueous working stocks are preferred over long-standing ones.

Analytical quantification in biological matrices generally targets the enzyme product rather than the inhibitor. Liquid chromatography with tandem mass spectrometry is the established route for 1-methylnicotinamide, with validated human plasma and urine methods reaching lower limits near 2 nanograms per millilitre. All of the above concerns laboratory characterisation of a research chemical. The material is intended for in vitro and preclinical research use only.