Signaling pathways intersecting with the p21-activated kinases (PAKs) play important roles in tumorigenesis and cancer progression. By recognizing that the limitations of FRAX1036 (1) were chiefly associated with the highly basic amine it contained, we devised a mitigation strategy to address several issues such as hERG activity. The 5-amino-1,3-dioxanyl moiety was identified as an effective means of reducing pK a and logP simultaneously. When positioned properly within the scaffold, this group conferred several benefits including potency, pharmacokinetics, and selectivity. Mouse xenograft PK/PD studies were carried out using an advanced compound, G-5555 (12), derived from this approach. These studies concluded that dose-dependent pathway modulation was achievable and paves the way for further in vivo investigations of PAK1 function in cancer and other diseases.

p21-activated kinase 1 (PAK1) has an important role in transducing signals in several oncogenic pathways. The concept of inhibiting this kinase has garnered significant interest over the past decade, particularly for targeting cancers associated with PAK1 amplification. Animal studies with the selective group I PAK (pan-PAK1, 2, 3) inhibitor G-5555 from the pyrido[2,3-d]pyrimidin-7-one class uncovered acute toxicity with a narrow therapeutic window. To attempt mitigating the toxicity, we introduced significant structural changes, culminating in the discovery of the potent pyridone side chain analogue G-9791. Mouse tolerability studies with this compound, other members of this series, and compounds from two structurally distinct classes revealed persistent toxicity and a correlation of minimum toxic concentrations and PAK1/2 mediated cellular potencies. Broad screening of selected PAK inhibitors revealed PAK1, 2, and 3 as the only overlapping targets. Our data suggest acute cardiovascular toxicity resulting from the inhibition of PAK2, which may be enhanced by PAK1 inhibition, and cautions against continued pursuit of pan-group I PAK inhibitors in drug discovery.

Inhibition of phosphoinositide 3-kinase (PI3K) signaling is an appealing approach to treat brain tumors, especially glioblastoma multiforme (GBM). We previously disclosed our successful approach to prospectively design potent and blood-brain barrier (BBB) penetrating PI3K inhibitors. The previously disclosed molecules were ultimately deemed not suitable for clinical development due to projected poor metabolic stability in humans. We, therefore, extended our studies to identify a BBB penetrating inhibitor of PI3K that was also projected to be metabolically stable in human. These efforts required identification of a distinct scaffold for PI3K inhibitors relative to our previous efforts and ultimately resulted in the identification of GDC-0084 (16). The discovery and preclinical characterization of this molecule are described within.

USP7 is a deubiquitinase implicated in destabilizing the tumor suppressor p53, and for this reason it has gained increasing attention as a potential oncology target for small molecule inhibitors. Herein we describe the biophysical, biochemical, and computational approaches that led to the identification of 4-(2-aminopyridin-3-yl)phenol compounds described by Kategaya ( Nature 2017 , 550 , 534 - 538 ) as specific inhibitors of USP7. Fragment based lead discovery (FBLD) by NMR combined with virtual screening and re-mining of biochemical high-throughput screening (HTS) hits led to the discovery of a series of ligands that bind in the "palm" region of the catalytic domain of USP7 and inhibit its catalytic activity. These ligands were then optimized by structure-based design to yield cell-active molecules with reasonable physical properties. This discovery process not only involved multiple techniques working in concert but also illustrated a unique way in which hits from orthogonal screening approaches complemented each other for lead identification.

Diverse biological roles for mitogen-activated protein kinase kinase kinase kinase 4 (MAP4K4) have necessitated the identification of potent inhibitors in order to study its function in various disease contexts. In particular, compounds that can be used to carry out such studies in vivo would be critical for elucidating the potential for therapeutic intervention. A structurebased design effort coupled with property-guided optimization directed at minimizing the ability of the inhibitors to cross into the CNS led to an advanced compound 13 (GNE-495) that showed excellent potency and good PK and was used to demonstrate in vivo efficacy in a retinal angiogenesis model recapitulating effects that were observed in the inducible Map4k4 knockout mice.