Design and Synthesis of Pyrrolo[2,3-d]pyrimidine-Derived Leucine-Rich Repeat Kinase 2 (LRRK2) Inhibitors Using a Checkpoint Kinase 1 (CHK1)-Derived Crystallographic Surrogate.

Inhibitors of leucine-rich repeat kinase 2 (LRRK2) and mutants, such as G2019S, have potential utility in Parkinson's disease treatment. Fragment hit-derived pyrrolo[2,3-d]pyrimidines underwent optimization using X-ray structures of LRRK2 kinase domain surrogates, based on checkpoint kinase 1 (CHK1) and a CHK1 10-point mutant. (2R)-2-Methylpyrrolidin-1-yl derivative 18 (LRRK2 G2019S cKi 0.7 nM, LE 0.66) was identified, with increased potency consistent with an X-ray structure of 18/CHK1 10-pt. mutant showing the 2-methyl substituent proximal to Ala147 (Ala2016 in LRRK2). Further structure-guided elaboration of 18 gave the 2-[(1,3-dimethyl-1H-pyrazol-4-yl)amino] derivative 32. Optimization of 32 afforded diastereomeric oxolan-3-yl derivatives 44 and 45, which demonstrated a favorable in vitro PK profile, although they displayed species disconnects in the in vivo PK profile, and a propensity for P-gp- and/or BCRP-mediated efflux in a mouse model. Compounds 44 and 45 demonstrated high potency and exquisite selectivity for LRRK2 and utility as chemical probes for the study of LRRK2 inhibition.

Design of Leucine-Rich Repeat Kinase 2 (LRRK2) Inhibitors Using a Crystallographic Surrogate Derived from Checkpoint Kinase 1 (CHK1).

Mutations in leucine-rich repeat kinase 2 (LRRK2), such as G2019S, are associated with an increased risk of developing Parkinson's disease. Surrogates for the LRRK2 kinase domain based on checkpoint kinase 1 (CHK1) mutants were designed, expressed in insect cells infected with baculovirus, purified, and crystallized. X-ray structures of the surrogates complexed with known LRRK2 inhibitors rationalized compound potency and selectivity. The CHK1 10-point mutant was preferred, following assessment of surrogate binding affinity with LRRK2 inhibitors. Fragment hit-derived arylpyrrolo[2,3-b]pyridine LRRK2 inhibitors underwent structure-guided optimization using this crystallographic surrogate. LRRK2-pSer935 HEK293 IC50 data for 22 were consistent with binding to Ala2016 in LRRK2 (equivalent to Ala147 in CHK1 10-point mutant structure). Compound 22 was shown to be potent, moderately selective, orally available, and brain-penetrant in wild-type mice, and confirmation of target engagement was demonstrated, with LRRK2-pSer935 IC50 values for 22 in mouse brain and kidney being 1.3 and 5 nM, respectively.

Application of Off-Rate Screening in the Identification of Novel Pan-Isoform Inhibitors of Pyruvate Dehydrogenase Kinase.

Libraries of nonpurified resorcinol amide derivatives were screened by surface plasmon resonance (SPR) to determine the binding dissociation constant (off-rate, kd) for compounds binding to the pyruvate dehydrogenase kinase (PDHK) enzyme. Parallel off-rate measurements against HSP90 and application of structure-based drug design enabled rapid hit to lead progression in a program to identify pan-isoform ATP-competitive inhibitors of PDHK. Lead optimization identified selective sub-100-nM inhibitors of the enzyme which significantly reduced phosphorylation of the E1α subunit in the PC3 cancer cell line in vitro.

Identification of novel, in vivo active Chk1 inhibitors utilizing structure guided drug design.

Chk1 kinase is a critical component of the DNA damage response checkpoint especially in cancer cells and targeting Chk1 is a potential therapeutic opportunity for potentiating the anti-tumor activity of DNA damaging chemotherapy drugs. Fragment elaboration by structure guided design was utilized to identify and develop a novel series of Chk1 inhibitors culminating in the identification of V158411, a potent ATP-competitive inhibitor of the Chk1 and Chk2 kinases. V158411 abrogated gemcitabine and camptothecin induced cell cycle checkpoints, resulting in the expected modulation of cell cycle proteins and increased cell death in cancer cells. V158411 potentiated the cytotoxicity of gemcitabine, cisplatin, SN38 and camptothecin in a variety of p53 deficient human tumor cell lines in vitro, p53 proficient cells were unaffected. In nude mice, V158411 showed minimal toxicity as a single agent and in combination with irinotecan. In tumor bearing animals, V158411 was detected at high levels in the tumor with a long elimination half-life; no pharmacologically significant in vivo drug-drug interactions with irinotecan were identified through analysis of the pharmacokinetic profiles. V158411 potentiated the anti-tumor activity of irinotecan in a variety of human colon tumor xenograft models without additional systemic toxicity. These results demonstrate the opportunity for combining V158411 with standard of care chemotherapeutic agents to potentiate the therapeutic efficacy of these agents without increasing their toxicity to normal cells. Thus, V158411 would warrant further clinical evaluation.

Leading the way: changing the focus from teaching to learning in large subjects with limited budgets.

To lead positive change in the teaching practice of teams that service large numbers of diverse students from multiple degree programs provides many challenges. The primary aim of this study was to provide a clear framework on which to plan the process of change that can be utilized by academic departments sector wide. Barriers to change were reduced by adapting and utilizing Kotter's principals of change specifically by creating a sense of urgency and defining a clear goal designed to address the problem. Changing attitudes involved training staff in new teaching and learning approaches and strategies, and creating a collaborative, supportive team-based teaching environment within which the planned changes could be implemented and evaluated. As a result senior academics are now directly involved in delivering sections of the face-to-face teaching in the new environment. Through promoting positive change we enabled deeper student engagement with the theoretical concepts delivered in lectures as evidenced by favorable student evaluations, feedback, and improved final exam results. A collaborative team-based approach that recognizes the importance of distributed leadership combined with a clearly articulated change management process were central to enabling academics to design, try, and evaluate the new teaching and learning practices. Our study demonstrates that a concerted focus on "change management" enabled teaching team members to adopt a major shift in the teaching and learning approach that resulted in measurable improvements in student learning.

14N quadrupole resonance and 1H T1 dispersion in the explosive RDX.

The explosive hexahydro-1,3,5-trinitro-s-triazine (CH2-N-NO2)3, commonly known as RDX, has been studied by 14N NQR and 1H NMR. NQR frequencies and relaxation times for the three ν+ and ν- lines of the ring 14N nuclei have been measured over the temperature range 230-330 K. The 1H NMR T1 dispersion has been measured for magnetic fields corresponding to the 1H NMR frequency range of 0-5.4 M Hz. The results have been interpreted as due to hindered rotation of the NO2 group about the N-NO2 bond with an activation energy close to 92 kJ mol(-1). Three dips in the 1H NMR dispersion near 120, 390 and 510 kHz are assigned to the ν0, ν- and ν+ transitions of the 14NO2 group. The temperature dependence of the inverse line-width parameters T2∗ of the three ν+ and ν- ring nitrogen transitions between 230 and 320 K can be explained by a distribution in the torsional oscillational amplitudes of the NO2 group about the N-NO2 bond at crystal defects whose values are consistent with the latter being mainly edge dislocations or impurities in the samples studied. Above 310 K, the 14N line widths are dominated by the rapid decrease in the spin-spin relaxation time T2 due to hindered rotation of the NO2 group. A consequence of this is that above this temperature, the 1H T1 values at the quadrupole dips are dominated by the spin mixing time between the 1H Zeeman levels and the combined 1H and 14N spin-spin levels.

Discovery and optimization of potent and selective functional antagonists of the human adenosine A2B receptor.

We herein report the discovery of a novel class of antagonists of the human adenosine A2B receptor. This low molecular weight scaffold has been optimized to offer derivatives with potential utility for the alleviation of conditions associated with this receptor subtype, such as nociception, diabetes, asthma and COPD. Furthermore, preliminary pharmacokinetic analysis has revealed compounds with profiles suitable for either inhaled or systemic routes of administration.

Stereoselective synthesis of 2,5-disubstituted-1,4-oxathiane S-oxides.

Beta-allyloxy and beta-propargyloxy tert-butyl sulfoxides undergo tandem sulfoxide eliminination-intramolecular sulfenic acid addition reactions to produce 1,4-oxathiane S-oxides.