Discovery of a series of ester-substituted NLRP3 inflammasome inhibitors.

The NLRP3 inflammasome is a component of the innate immune system involved in the production of proinflammatory cytokines. Aberrant activation by a wide range of exogenous and endogenous signals can lead to chronic, low-grade inflammation. It has attracted a great deal of interest as a drug target due to the association with diseases of large unmet medical need such as Alzheimer's disease, Parkinson's disease, arthritis, and cancer. To date, no drugs specifically targeting inhibition of the NLRP3 inflammasome have been approved. In this work, we used the known NLRP3 inflammasome inhibitor CP-456,773 (aka CRID3 or MCC 950) as our starting point and undertook a Structure-Activity Relationship (SAR) analysis and subsequent scaffold-hopping exercise. This resulted in the rational design of a series of novel ester-substituted urea compounds that are highly potent and selective NLRP3 inflammasome inhibitors, as exemplified by compounds 44 and 45. It is hypothesized that the ester moiety acts as a highly permeable delivery vehicle and is subsequently hydrolyzed to the carboxylic acid active species by carboxylesterase enzymes. These molecules are greatly differentiated from the state-of-the-art and offer potential in the treatment of NLRP3-driven diseases, particularly where tissue penetration is required.

N-Arylation of Diketopyrrolopyrroles with Aryl Triflates.

A new methodology for the double N-arylation of diketopyrrolopyrroles with aryl triflates has been developed. It is now possible to prepare diketopyrrolopyrroles bearing N-substituents derived from naphthalene, anthracene and coumarin in two steps from commercially available phenols. This represents the first time arenes lacking strong electron-withdrawing groups were inserted onto lactamic nitrogen atoms via arylation. The ability to incorporate heretofore unprecedented substituents translates to increased modulation of the resulting photophysical properties such as switching-on/off solvatofluorochromism. TD-DFT calculations have been performed to explore the nature of the relevant excited states. This new synthetic method made it possible to elucidate the influence of such substituents on the absorption and emission properties of tetraaryl substituted diketopyrrolopyrroles.

5-Keto-3-cyano-2,4-diaminothiophenes as selective maternal embryonic leucine zipper kinase inhibitors.

Maternal embryonic leucine zipper kinase (MELK) is involved in several key cellular processes and displays increased levels of expression in numerous cancer classes (colon, breast, brain, ovary, prostate and lung). Although no selective MELK inhibitors have yet been approved, increasing evidence suggest that inhibition of MELK would constitute a promising approach for cancer therapy. A weak high-throughput screening hit (17, IC50 ≈ 5 µM) with lead-like properties was optimized for MELK inhibition. The early identification of a plausible binding mode by molecular modeling offered guidance in the choice of modifications towards compound 52 which displayed a 98 nM IC50. A good selectivity profile was achieved for a representative member of the series (29) in a 486 protein kinase panel. Future elaboration of 52 has the potential to deliver compounds for further development with chemotherapeutic aims.

A novel, dual pan-PIM/FLT3 inhibitor SEL24 exhibits broad therapeutic potential in acute myeloid leukemia.

Fms-like tyrosine kinase 3 internal tandem duplication (FLT3-ITD) is one of the most common genetic lesions in acute myeloid leukemia patients (AML). Although FLT3 tyrosine kinase inhibitors initially exhibit clinical activity, resistance to treatment inevitably occurs within months. PIM kinases are thought to be major drivers of the resistance phenotype and their inhibition in relapsed samples restores cell sensitivity to FLT3 inhibitors. Thus, simultaneous PIM and FLT3 inhibition represents a promising strategy in AML therapy. For such reasons, we have developed SEL24-B489 - a potent, dual PIM and FLT3-ITD inhibitor. SEL24-B489 exhibited significantly broader on-target activity in AML cell lines and primary AML blasts than selective FLT3-ITD or PIM inhibitors. SEL24-B489 also demonstrated marked activity in cells bearing FLT3 tyrosine kinase domain (TKD) mutations that lead to FLT3 inhibitor resistance. Moreover, SEL24-B489 inhibited the growth of a broad panel of AML cell lines in xenograft models with a clear pharmacodynamic-pharmacokinetic relationship. Taken together, our data highlight the unique dual activity of the SEL24-B489 that abrogates the activity of signaling circuits involved in proliferation, inhibition of apoptosis and protein translation/metabolism. These results underscore the therapeutic potential of the dual PIM/FLT3-ITD inhibitor for the treatment of AML.

Electron ionization and electrospray mass spectra of diaryl-substituted enaminoketones and their thio analogs.

Electron ionization (EI) and positive electrospray ionization (ESI) mass spectra of selected diaryl enaminoketones and enaminothiones have been studied. In the EI mass spectra of both classes of compound, molecular ion peaks are accompanied by the peaks corresponding to the [M-H](+) ions. The formation of these ions can be rationalized by a cyclization reaction resulting in the formation of the respective isoxazolium and isothiazolium cations. Under positive ESI conditions, in the spectra recorded for the enaminoketones peaks corresponding to the [M+H](+), [M+Na](+) and [2M+Na](+) ions appeared, while in the spectra recorded for the enaminothiones, peaks corresponding to the [M-H](+) ions were dominant. These ions are most likely formed by oxidation of the neutral enaminothione molecules on the surface of the positively charged stainless steel capillary in the ESI ion source (anodic oxidation).

Structural studies on aryl-substituted enaminoketones and their thio analogues. Part II. Experimental and DFT calculated carbon-carbon coupling constants, (n)J(CC)'s (n = 1-3).

Spin-spin carbon-carbon coupling constants across one, two and three bonds, J(CC), have been measured for a series of aryl-substituted Z-s-Z-s-E enaminoketones and their thio analogues. As a result, a large set, altogether 178, of J(CC)s has been obtained. It consists of 82 couplings across one bond, 31 couplings across two bonds and 65 couplings across three bonds. Independently, the DFT calculations at the B3PW91/6-311++G(d,p)//B3PW91/6-311++G(d,p) level yielded a set of theoretical J(CC) values. A comparison of these two sets of data gave an excellent linear correlation with parameters a and b close to ideal; a = 0.9978 which is not far from unity and b = 0.22 Hz which is close to zero. The (1)J(CC) couplings determined for the crucial fragment of the molecules, i.e. -C=C-C=O (or -C=C-C=S), are: (1)J(C=C) approximately 68 Hz (67 Hz) and (1)J(C-C) = 60.5 Hz (60.0 Hz). The corresponding couplings found for the Z-s-Z-s-E isomer of the parent enaminoketone, 4-methylamino-but-3-en-2-one are 64.1 and 59.3 Hz, respectively. The most sensitive towards substitution of the oxygen atom by sulfur are two-bond couplings between the alpha-vinylic and aromatic C(ipso) carbon atoms, which attain 12 Hz in the enaminoketone derivatives and decrease to 5 Hz in their thio analogues.

Structural studies on aryl-substituted enaminoketones and their thio analogues. Part I. Analysis of high-resolution (1)H, (13)C NMR and (13)C CP MAS spectra combined with GIAO-DFT calculations.

A series of aryl-substituted enaminoketones and their thio analogues in CDCl(3) solution and in the solid state were studied by the use of high-resolution (1)H and (13)C as well as (13)C cross polarization magic angle spinning (CP MAS) NMR spectra in combination with gauge including atomic orbitals-density functional theory (GIAO-DFT) calculations performed at the B3PW91/6-311 + + G(d,p) level of theory using the B3PW91/6-311 + + G(d,p)-optimized geometries. The analysis of the (13)C NMR spectra in solution was done by using the Incredible Natural Abundance DoublE QUAntum Transfer Experiment (INADEQUATE) technique, whereas trends observed in the (13)C shielding constants, calculated for the compounds studied, were a great help in assigning most of the signals in the (13)C CP MAS NMR spectra. It was established on the basis of the experimental and theoretical NMR data that both groups of compounds exist in the form of Z-s-Z-s-E isomers in CDCl(3) solution as well as in the solid state, with the NH hydrogen atom involved in intramolecular hydrogen bonding. This conclusion is in agreement with the fact that some of the compounds studied reveal liquid-crystalline properties. Three-bond H, H and C, H coupling constants measured in solution played a crucial role in the structure elucidation.

Synthesis and biological activity of N(alpha)-[4-[N-[(3,4-dihydro-2-methyl-4-oxo-6-quinazolinyl)methyl]-N-propargylamino]phenylacetyl]-L-glutamic acid.

2-Deamino-2-methyl-N10-propargyl-5,8-dideazafolic acid (ICI 198583) is a potent inhibitor of thymidylate synthase. Its analogue, N(alpha)-[4-[N-[(3,4-dihydro-2-methyl-4-oxo-6-quinazolinyl)methyl]-N-propargylamino]phenylacetyl]-L-glutamic acid, containing p-aminophenylacetic acid residue substituting p-aminobenzoic acid residue, was synthesized. The new analogue exhibited a moderately potent thymidylate synthase inhibition, of linear mixed type vs. the cofactor, N(5,10)-methylenetetrahydrofolate. The Ki value of 0.34 microM, determined with a purified recombinant rat hepatoma enzyme, was about 30-fold higher than that reported for inhibition of thymidylate synthase from mouse leukemia L1210 cells by ICI 198583 (Hughes et al., 1990, J. Med. Chem. 33, 3060). Growth of mouse leukemia L5178Y cells was inhibited by the analogue (IC50 = 1.26 mM) 180-fold weaker than by ICI 198583 (IC50 = 6.9 microM).