KRAS and BRAF activating mutations drive tumorigenesis through constitutive activation of the MAPK pathway. As these tumours represent an area of high unmet medical need, multiple allosteric MEK inhibitors, which inhibit MAPK signalling in both genotypes, are being tested in clinical trials. Impressive single-agent activity in BRAF-mutant melanoma has been observed; however, efficacy has been far less robust in KRAS-mutant disease. Here we show that, owing to distinct mechanisms regulating MEK activation in KRAS- versus BRAF-driven tumours, different mechanisms of inhibition are required for optimal antitumour activity in each genotype. Structural and functional analysis illustrates that MEK inhibitors with superior efficacy in KRAS-driven tumours (GDC-0623 and G-573, the former currently in phase I clinical trials) form a strong hydrogen-bond interaction with S212 in MEK that is critical for blocking MEK feedback phosphorylation by wild-type RAF. Conversely, potent inhibition of active, phosphorylated MEK is required for strong inhibition of the MAPK pathway in BRAF-mutant tumours, resulting in superior efficacy in this genotype with GDC-0973 (also known as cobimetinib), a MEK inhibitor currently in phase III clinical trials. Our study highlights that differences in the activation state of MEK in KRAS-mutant tumours versus BRAF-mutant tumours can be exploited through the design of inhibitors that uniquely target these distinct activation states of MEK. These inhibitors are currently being evaluated in clinical trials to determine whether improvements in therapeutic index within KRAS versus BRAF preclinical models translate to improved clinical responses in patients.

The mild and selective hydrolysis of esters can often be crucial in the sequence toward a target molecule and is, therefore, an important objective in contemporary organic synthesis. Although several methods exist to accomplish this task in certain cases, a mild, generally applicable protocol remains absent. Frequent problems encountered include the concurrent hydrolysis of other ester groups present within the molecule under scrutiny, epimerization of stereocenters, and elimination reactions induced by the often basic conditions employed. Herein we report a new and selective method for the hydrolysis of esters under extremely mild conditions that avoid such side reactions and lead to high yields of the corresponding carboxylic acids.It was during our campaign toward thiostrepton, [1] a highly complex thiopeptide antibiotic, that we had the opportunity to search for such a method. Our sensitive intermediates proved too fragile to tolerate standard ester hydrolysis conditions. We finally came upon Me 3 SnOH, which had been previously employed by Mascaretti and co-workers [2] to cleave phenacyl ester anchored amino acids and peptides from a polystyrene resin and to hydrolyze methyl and isopropyl phenylacetate to give the corresponding acids in high yield. To our knowledge, these are the only examples in which Me 3 SnOH has been previously used to carry out hydrolytic ruptures of esters.[3] As shown in Table 1, this reagent proved extremely useful to us in attaining the highyielding and selective hydrolysis of methyl esters within the sensitive substrates 1-4, which were encountered en route to thiostrepton. These remarkable results prompted a secondphase investigation in which we attempted to determine systematically the generality and scope of this protocol, which involved heating the substrate with 1-10 equivalents of[*] Prof.

TYK2 is a JAK family protein tyrosine kinase activated in response to multiple cytokines, including type I IFNs, IL-6, IL-10, IL-12, and IL-23. Extensive studies of mice that lack TYK2 expression indicate that the IFN-α, IL-12, and IL-23 pathways, but not the IL-6 or IL-10 pathways, are compromised. In contrast, there have been few studies of the role of TYK2 in primary human cells. A genetic mutation at the tyk2 locus that results in a lack of TYK2 protein in a single human patient has been linked to defects in the IFN-α, IL-6, IL-10, IL-12, and IL-23 pathways, suggesting a broad role for TYK2 protein in human cytokine responses. In this article, we have used a panel of novel potent TYK2 small-molecule inhibitors with varying degrees of selectivity against other JAK kinases to address the requirement for TYK2 catalytic activity in cytokine pathways in primary human cells. Our results indicate that the biological processes that require TYK2 catalytic function in humans are restricted to the IL-12 and IL-23 pathways, and suggest that inhibition of TYK2 catalytic activity may be an efficacious approach for the treatment of select autoimmune diseases without broad immunosuppression.

The well accepted "free drug hypothesis" for small-molecule drugs assumes that only the free (unbound) drug concentration at the therapeutic target can elicit a pharmacologic effect. Unbound (free) drug concentrations in plasma are readily measurable and are often used as surrogates for the drug concentrations at the site of pharmacologic action in pharmacokinetic-pharmacodynamic analysis and clinical dose projection in drug discovery. Furthermore, for permeable compounds at pharmacokinetic steady state, the free drug concentration in tissue is likely a close approximation of that in plasma; however, several factors can create and maintain disequilibrium between the free drug concentration in plasma and tissue, leading to free drug concentration asymmetry. These factors include drug uptake and extrusion mechanisms involving the uptake and efflux drug transporters, intracellular biotransformation of prodrugs, membrane receptor-mediated uptake of antibody-drug conjugates, pH gradients, unique distribution properties (covalent binders, nanoparticles), and local drug delivery (e.g., inhalation). The impact of these factors on the free drug concentrations in tissues can be represented by K p,uu , the ratio of free drug concentration between tissue and plasma at steady state. This review focuses on situations in which free drug concentrations in tissues may differ from those in plasma (e.g., K p,uu > or <1) and discusses the limitations of the surrogate approach of using plasmafree drug concentration to predict free drug concentrations in tissue. This is an important consideration for novel therapeutic modalities since systemic exposure as a driver of pharmacologic effects may provide limited value in guiding compound optimization, selection, and advancement. Ultimately, a deeper understanding of the relationship between free drug concentrations in plasma and tissues is needed.

The first phase of the total synthesis of thiostrepton (1), a highly complex thiopeptide antibiotic, is described. After a brief introduction to the target molecule and its structural motifs, it is shown that retrosynthetic analysis of thiostrepton reveals compounds 23, 24, 26, 28, and 29 as potential key building blocks for the projected total synthesis. Concise and stereoselective constructions of all these intermediates are then described. The synthesis of the dehydropiperidine core 28 was based on a biosynthetically inspired aza-Diels-Alder dimerization of an appropriate azadiene system, an approach that was initially plagued with several problems which were, however, resolved satisfactorily by systematic investigations. The quinaldic acid fragment 24 and the thiazoline-thiazole segment 26 were synthesized by a series of reactions that included asymmetric and other stereoselective processes. The dehydroalanine tail precursor 23 and the alanine equivalent 29 were also prepared from the appropriate amino acids. Finally, a method was developed for the direct coupling of the labile dehydropiperidine key building block 28 to the more advanced and stable peptide intermediate 27 through capture with the highly reactive alanine equivalent 67 under conditions that avoided the initially encountered destructive ring contraction process.

Nicotinamide phosphoribosyltransferase (Nampt) is a promising anticancer target. Virtual screening identified a thiourea analogue, compound 5, as a novel highly potent Nampt inhibitor. Guided by the cocrystal structure of 5, SAR exploration revealed that the corresponding urea compound 7 exhibited similar potency with an improved solubility profile. These studies also indicated that a 3-pyridyl group was the preferred substituent at one inhibitor terminus and also identified a urea moiety as the optimal linker to the remainder of the inhibitor structure. Further SAR optimization of the other inhibitor terminus ultimately yielded compound 50 as a urea-containing Nampt inhibitor which exhibited excellent biochemical and cellular potency (enzyme IC50 = 0.007 μM; A2780 IC50 = 0.032 μM). Compound 50 also showed excellent in vivo antitumor efficacy when dosed orally in an A2780 ovarian tumor xenograft model (TGI of 97% was observed on day 17).

A therapeutic rationale is proposed for the treatment of inflammatory diseases, such as rheumatoid arthritis (RA), by specific targeting of the JAK1 pathway. Examination of the preferred binding conformation of clinically effective, pan-JAK inhibitor 1 led to identification of a novel, tricyclic hinge binding scaffold 3. Exploration of SAR through a series of cycloamino and cycloalkylamino analogues demonstrated this template to be highly tolerant of substitution, with a predisposition to moderate selectivity for the JAK1 isoform over JAK2. This study culminated in the identification of subnanomolar JAK1 inhibitors such as 22 and 49, having excellent cell potency, good rat pharmacokinetic characteristics, and excellent kinase selectivity. Determination of the binding modes of the series in JAK1 and JAK2 by X-ray crystallography supported the design of analogues to enhance affinity and selectivity.

The mild and selective hydrolysis of esters can often be crucial in the sequence toward a target molecule and is, therefore, an important objective in contemporary organic synthesis. Although several methods exist to accomplish this task in certain cases, a mild, generally applicable protocol remains absent. Frequent problems encountered include the concurrent hydrolysis of other ester groups present within the molecule under scrutiny, epimerization of stereocenters, and elimination reactions induced by the often basic conditions employed. Herein we report a new and selective method for the hydrolysis of esters under extremely mild conditions that avoid such side reactions and lead to high yields of the corresponding carboxylic acids.It was during our campaign toward thiostrepton, [1] a highly complex thiopeptide antibiotic, that we had the opportunity to search for such a method. Our sensitive intermediates proved too fragile to tolerate standard ester hydrolysis conditions. We finally came upon Me 3 SnOH, which had been previously employed by Mascaretti and co-workers [2] to cleave phenacyl ester anchored amino acids and peptides from a polystyrene resin and to hydrolyze methyl and isopropyl phenylacetate to give the corresponding acids in high yield. To our knowledge, these are the only examples in which Me 3 SnOH has been previously used to carry out hydrolytic ruptures of esters.[3] As shown in Table 1, this reagent proved extremely useful to us in attaining the highyielding and selective hydrolysis of methyl esters within the sensitive substrates 1-4, which were encountered en route to thiostrepton. These remarkable results prompted a secondphase investigation in which we attempted to determine systematically the generality and scope of this protocol, which involved heating the substrate with 1-10 equivalents of[*] Prof.