Ina Dix

Discovery of Tetrahydropyrazolopyrimidine Carboxamide Derivatives As Potent and Orally Active Antitubercular Agents

Tetrahydropyrazolo[1,5-a]pyrimidine scaffold was identified as a hit series from a Mycobacterium tuberculosis (Mtb) whole cell high through-put screening (HTS) campaign. A series of derivatives of this class were synthesized to evaluate their structure-activity relationship (SAR) and structure-property relationship (SPR). Compound 9 had a promising in vivo DMPK profile in mouse and exhibited potent in vivo activity in a mouse efficacy model, achieving a reduction of 3.5 log CFU of Mtb after oral administration to infected mice once a day at 100 mg/kg for 28 days. Thus, compound 9 is a potential candidate for inclusion in combination therapies for both drug-sensitive and drug-resistant TB.

Design and Development of a Cyclic Decapeptide Scaffold with Suitable Properties for Bioavailability and Oral Exposure

Permeability and oral bioavailability of macrocyclic peptides still represent difficult challenges in drug discovery. Despite the recognized potential of macrocyclic peptides as therapeutics, their use is still restricted to extracellular targets and intravenous administration. Indeed, macrocyclic peptides generally suffer from limited proteolytic stability, high clearance, and poor membrane permeability, and this leads to the absence of systemic exposure after oral administration. To overcome these limitations, we started to investigate the development of a general cyclic decapeptide scaffold that possesses ideal features for cell permeability and oral exposure. On the basis of a rigid hairpin structure, the scaffold design aimed to decrease the overall polarity of the compound, thereby limiting the energetic cost of NH desolvation and the entropy penalty during cell penetration. The results of this study also demonstrate the importance of rigidity for the β‐turn design regarding clearance. To stabilize the scaffold in the desired β‐hairpin conformation, the introduction of d‐proline at the i+1 turn position proved to be beneficial for both permeability and clearance. As a result, cyclopeptide decamers with unprecedented high values for oral bioavailability and exposure are reported herein. NMR spectroscopy conformation and dynamic analysis confirmed, for selected examples, the rigidity of the scaffold and the presence of transannular hydrogen bonds in polar and apolar environments. Furthermore, we showed, for one compound, that its transition from a polar environment to an apolar one was accompanied by an increased molecular motion, revealing an entropy contribution to membrane permeation.

Lead Optimization of Spiropyrazolopyridones: A New and Potent Class of Dengue Virus Inhibitors

Spiropyrazolopyridone 1 was identified, as a novel dengue virus (DENV) inhibitor, from a DENV serotype 2 (DENV-2) high-throughput phenotypic screen. As a general trend within this chemical class, chiral resolution of the racemate revealed that R enantiomer was significantly more potent than the S. Cell-based lead optimization of the spiropyrazolopyridones focusing on improving the physicochemical properties is described. As a result, an optimal compound 14a, with balanced in vitro potency and pharmacokinetic profile, achieved about 1.9 log viremia reduction at 3 × 50 mg/kg (bid) or 3 × 100 mg/kg (QD) oral doses in the dengue in vivo mouse efficacy model.

Optimization of 3-Pyrimidin-4-yl-oxazolidin-2-ones as Allosteric and Mutant Specific Inhibitors of IDH1

High throughput screening and subsequent hit validation identified 4-isopropyl-3-(2-((1-phenylethyl)amino)pyrimidin-4-yl)oxazolidin-2-one as a potent inhibitor of IDH1R132H. Synthesis of the four separate stereoisomers identified the (S,S)-diastereomer (IDH125, 1f) as the most potent isomer. This also showed reasonable cellular activity and excellent selectivity vs IDH1wt. Initial structure-activity relationship exploration identified the key tolerances and potential for optimization. X-ray crystallography identified a functionally relevant allosteric binding site amenable to inhibitors, which can penetrate the blood-brain barrier, and aided rational optimization. Potency improvement and modulation of the physicochemical properties identified (S,S)-oxazolidinone IDH889 (5x) with good exposure and 2-HG inhibitory activity in a mutant IDH1 xenograft mouse model.

Design and Discovery of N-(2-Methyl-5′-morpholino-6′-((tetrahydro-2H-pyran-4-yl)oxy)-[3,3′-bipyridin]-5-yl)-3-(trifluoromethyl)benzamide (RAF709): A Potent, Selective, and Efficacious RAF Inhibitor Targeting RAS Mutant Cancers

RAS oncogenes have been implicated in >30% of human cancers, all representing high unmet medical need. The exquisite dependency on CRAF kinase in KRAS mutant tumors has been established in genetically engineered mouse models and human tumor cells. To date, many small molecule approaches are under investigation to target CRAF, yet kinase-selective and cellular potent inhibitors remain challenging to identify. Herein, we describe 14 (RAF709) [ Aversa , Biaryl amide compounds as kinase inhibitors and their preparation . WO 2014151616, 2014 ], a selective B/C RAF inhibitor, which was developed through a hypothesis-driven approach focusing on drug-like properties. A key challenge encountered in the medicinal chemistry campaign was maintaining a balance between good solubility and potent cellular activity (suppression of pMEK and proliferation) in KRAS mutant tumor cell lines. We investigated the small molecule crystal structure of lead molecule 7 and hypothesized that disruption of the crystal packing would improve solubility, which led to a change from N-methylpyridone to a tetrahydropyranyl oxy-pyridine derivative. 14 proved to be soluble, kinase selective, and efficacious in a KRAS mutant xenograft model.

Construction of a 3D-shaped, natural product like fragment library by fragmentation and diversification of natural products

A fragment library consisting of 3D-shaped, natural product-like fragments was assembled. Library construction was mainly performed by natural product degradation and natural product diversification reactions and was complemented by the identification of 3D-shaped, natural product like fragments available from commercial sources. In addition, during the course of these studies, novel rearrangements were discovered for Massarigenin C and Cytochalasin E. The obtained fragment library has an excellent 3D-shape and natural product likeness, covering a novel, unexplored and underrepresented chemical space in fragment based drug discovery (FBDD).

Discovery of 2-oxopiperazine dengue inhibitors by scaffold morphing of a phenotypic high-throughput screening hit

A series of 2-oxopiperazine derivatives were designed from the pyrrolopiperazinone cell-based screening hit 4 as a dengue virus inhibitor. Systematic investigation of the structure-activity relationship (SAR) around the piperazinone ring led to the identification of compound (S)-29, which exhibited potent anti-dengue activity in the cell-based assay across all four dengue serotypes with EC50<0.1μM. Cross-resistant analysis confirmed that the virus NS4B protein remained the target of the new oxopiperazine analogs obtained via scaffold morphing from the HTS hit 4.

Bioinspired co-crystals of Imatinib providing enhanced kinetic solubility

Graphical abstract Figure. No caption available. Abstract Realizing the full potential of co‐crystals enhanced kinetic solubility demands a comprehensive understanding of the mechanisms of dissolution, phase conversion, nucleation and crystal growth, and of the complex interplay between the active pharmaceutical ingredient (API), the coformer and co‐existing forms in aqueous media. One blueprint provided by nature to keep poorly water‐soluble bases in solution is the complexation with phenolic acids. Consequently, we followed a bioinspired strategy for the engineering of co‐crystals of a poorly water‐soluble molecule – Imatinib – with a phenolic acid, syringic acid (SYA). The dynamics of dissolution and solution‐mediated phase transformations were monitored by Nuclear Magnetic Resonance (NMR) spectroscopy, providing mechanistic insights into the 60 fold‐increased long lasting concentrations achieved by the syringate co‐crystals as compared to Imatinib base and Imatinib mesylate. This lasting effect was linked to SYA’s ability to delay the formation and nucleation of Imatinib hydrate – the thermodynamically stable form in aqueous media – through a metastable association of SYA with Imatinib in solution. Results from permeability studies evidenced that SYA did not impact Imatinib’s permeability across membranes while suggesting improved bioavailability through higher kinetic solubility at the biological barriers. These results reflect that some degree of hydrophobicity of the coformer might be key to extend the kinetic solubility of co‐crystals with hydrophobic APIs. Understanding how kinetic supersaturation can be shaped by the selection of an interactive coformer may help achieving the needed performance of new forms of poorly water‐soluble, slowly dissolving APIs.

GuideX: a systematic approach to managing and archiving crystallization data

In the pharmaceutical industry the threedimensional structure of a molecule plays a critical role in drug development and the investigation of structure-activity-relationships (SAR). The classical and still most reliable way to obtain those 3D models is single crystal x-ray structure analysis. The bottleneck of this technique, however, is the necessity to grow single crystals suitable for the analysis. This is especially challenging when only a few milligrams of the compound of interest are available and automation or large scale screens are not possible. Despite many efforts to understand the theory behind crystal growth the method of choice in practice is often enough trial and error mixed with experience and gut feeling. Hence, it is difficult to share any of this knowledge within the broader crystallographic community. [1], [2] The attention the recently published “crystal sponge method” has received, however, emphasizes once again the need for novel, nonconventional approaches.[3] Our own idea to reduce the bottleneck is based on the hypothesis that the crystallization propensity is dominated by the intrinsic properties of the individual compound. It aims at a statistical and systematic analysis of our in-house crystallization records to derive predictive classification tools and maps of crystallization propensity in descriptor space. To be able to extract significant information from our data, however, it was first necessary to develop a highly systematic and ordered procedure for the crystallization process which allows tracking of all activities, parameters and results. This procedure will be presented. The associated data of more than 300 crystallization requests per year (from the experimental parameters to the results of each trial) is stored electronically in a specifically designed Oracle database. The data can be entered on the fly simultaneously to the experimental work at the hood, the visual inspection at the microscope and the crystal quality check on the diffractometer via GuideX, an in-house touch screen application which is directly linked to the above mentioned database. Since we use defined sets of qualifiers to describe all observations from solubility behavior to crystallization results we are also able to derive fingerprints for each compound which might lead to a new kind of classification. First results and findings will be presented and will be the foundation for further systematic investigations. We consider our GuideX “crystallization data management system” the first step on the way to deriving structure based rules which then can guide the crystallization protocol for unknown organic compounds and thereby significantly reduce the time and material spent on this process.