Sergio G. Durón

Rescue of fragile X syndrome phenotypes in Fmr1 KO mice by the small-molecule PAK inhibitor FRAX486

Fragile X syndrome (FXS) is the most common inherited form of autism and intellectual disability and is caused by the silencing of a single gene, fragile X mental retardation 1 (Fmr1). The Fmr1 KO mouse displays phenotypes similar to symptoms in the human condition—including hyperactivity, repetitive behaviors, and seizures—as well as analogous abnormalities in the density of dendritic spines. Here we take a hypothesis-driven, mechanism-based approach to the search for an effective therapy for FXS. We hypothesize that a treatment that rescues the dendritic spine defect in Fmr1 KO mice may also ameliorate autism-like behavioral symptoms. Thus, we targeted a protein that regulates spines through modulation of actin cytoskeleton dynamics: p21-activated kinase (PAK). Our results demonstrate that a potent small molecule inhibitor of group I PAKs reverses dendritic spine phenotypes in Fmr1 KO mice. Moreover, this PAK inhibitor—which we call FRAX486—also rescues seizures and behavioral abnormalities such as hyperactivity and repetitive movements, thereby supporting the hypothesis that a drug treatment that reverses the spine abnormalities can also treat neurological and behavioral symptoms. Finally, a single administration of FRAX486 is sufficient to rescue all of these phenotypes in adult Fmr1 KO mice, demonstrating the potential for rapid, postdiagnostic therapy in adults with FXS.

Reactivity-based one-pot total synthesis of fucose GM1 oligosaccharide: A sialylated antigenic epitope of small-cell lung cancer

The total synthesis of the sialic acid-containing antigenic epitope fucose GM1 (Fuc-GM1) by an improved reactivity-based one-pot synthetic strategy is reported. Based on a thioglycoside reactivity database, three saccharide building blocks, 3, 4, and 5, were designed and prepared to incorporate a descending order of reactivity toward thiophilic activation. Using the reactivity-based one-pot synthetic method, the fully protected Fuc-GM1 glycoside 2 was furnished in a facile manner, which was globally deprotected to give the Fuc-GM1 glycoside 1. In addition, using the promoter system 1-(benzensulfinyl)piperidine/trifluoromethanesulfonic anhydride, the product yield was improved and the reaction time was reduced in comparison with the N-iodosuccinimide/trifluoromethanesulfonic acid- and dimethyl (thiomethyl) sulfonium trifluoromethanesulfonate-promoted systems.

p21-Activated Kinase 1 Is Required for Efficient Tumor Formation and Progression in a Ras-Mediated Skin Cancer Model

The RAS genes are the most commonly mutated oncogenes in human cancer and present a particular therapeutic dilemma, as direct targeting of Ras proteins by small molecules has proved difficult. Signaling pathways downstream of Ras, in particular Raf/Mek/Erk and PI3K/Akt/mTOR, are dominated by lipid and protein kinases that provide attractive alternate targets in Ras-driven tumors. As p21-activated kinase 1 (Pak1) has been shown to regulate both these signaling pathways and is itself upregulated in many human cancers, we assessed the role of Pak1 in Ras-driven skin cancer. In human squamous cell carcinoma (SCC), we found a strong positive correlation between advanced stage and grade and PAK1 expression. Using a mouse model of Kras-driven SCC, we showed that deletion of the mouse Pak1 gene led to markedly decreased tumorigenesis and progression, accompanied by near total loss of Erk and Akt activity. Treatment of Kras(G12D) mice with either of two distinct small molecule Pak inhibitors (PF3758309 and FRAX597) caused tumor regression and loss of Erk and Akt activity. Tumor regression was also seen in mice treated with a specific Mek inhibitor, but not with an Akt inhibitor. These findings establish Pak1 as a new target in KRAS-driven tumors and suggest a mechanism of action through the Erk, but not the Akt, signaling pathway.

PAKs inhibitors ameliorate schizophrenia-associated dendritic spine deterioration in vitro and in vivo during late adolescence.

Significance Drug discovery in psychiatry has been limited to chemical modifications of compounds originally discovered serendipitously. Therefore, more mechanism-oriented strategies of drug discovery for mental disorders are awaited. Schizophrenia is a devastating mental disorder with synaptic disconnectivity involved in its pathophysiology. In this study, we studied a biological pathway underlying synaptic disturbance and examined whether p21-activated kinase inhibitors ameliorate the pathology in vitro and in vivo. The beneficial effects of these inhibitors reported here may provide us with an opportunity for drug discovery in major mental illnesses with synaptic disturbance. Drug discovery in psychiatry has been limited to chemical modifications of compounds originally discovered serendipitously. Therefore, more mechanism-oriented strategies of drug discovery for mental disorders are awaited. Schizophrenia is a devastating mental disorder with synaptic disconnectivity involved in its pathophysiology. Reduction in the dendritic spine density is a major alteration that has been reproducibly reported in the cerebral cortex of patients with schizophrenia. Disrupted-in-Schizophrenia-1 (DISC1), a factor that influences endophenotypes underlying schizophrenia and several other neuropsychiatric disorders, has a regulatory role in the postsynaptic density in association with the NMDA-type glutamate receptor, Kalirin-7, and Rac1. Prolonged knockdown of DISC1 leads to synaptic deterioration, reminiscent of the synaptic pathology of schizophrenia. Thus, we tested the effects of novel inhibitors to p21-activated kinases (PAKs), major targets of Rac1, on synaptic deterioration elicited by knockdown expression of DISC1. These compounds not only significantly ameliorated the synaptic deterioration triggered by DISC1 knockdown but also partially reversed the size of deteriorated synapses in culture. One of these PAK inhibitors prevented progressive synaptic deterioration in adolescence as shown by in vivo two-photon imaging and ameliorated a behavioral deficit in prepulse inhibition in adulthood in a DISC1 knockdown mouse model. The efficacy of PAK inhibitors may have implications in drug discovery for schizophrenia and related neuropsychiatric disorders in general.

Small molecule inhibition of group I p21-activated kinases in breast cancer induces apoptosis and potentiates the activity of microtubule stabilizing agents

IntroductionBreast cancer, the most common cause of cancer-related deaths worldwide among women, is a molecularly and clinically heterogeneous disease. Extensive genetic and epigenetic profiling of breast tumors has recently revealed novel putative driver genes, including p21-activated kinase (PAK)1. PAK1 is a serine/threonine kinase downstream of small GTP-binding proteins, Rac1 and Cdc42, and is an integral component of growth factor signaling networks and cellular functions fundamental to tumorigenesis.MethodsPAK1 dysregulation (copy number gain, mRNA and protein expression) was evaluated in two cohorts of breast cancer tissues (n = 980 and 1,108). A novel small molecule inhibitor, FRAX1036, and RNA interference were used to examine PAK1 loss of function and combination with docetaxel in vitro. Mechanism of action for the therapeutic combination, both cellular and molecular, was assessed via time-lapse microscopy and immunoblotting.ResultsWe demonstrate that focal genomic amplification and overexpression of PAK1 are associated with poor clinical outcome in the luminal subtype of breast cancer (P = 1.29 × 10−4 and P = 0.015, respectively). Given the role for PAK1 in regulating cytoskeletal organization, we hypothesized that combination of PAK1 inhibition with taxane treatment could be combined to further interfere with microtubule dynamics and cell survival. Consistent with this, administration of docetaxel with either a novel small molecule inhibitor of group I PAKs, FRAX1036, or PAK1 small interfering RNA oligonucleotides dramatically altered signaling to cytoskeletal-associated proteins, such as stathmin, and induced microtubule disorganization and cellular apoptosis. Live-cell imaging revealed that the duration of mitotic arrest mediated by docetaxel was significantly reduced in the presence of FRAX1036, and this was associated with increased kinetics of apoptosis.ConclusionsTaken together, these findings further support PAK1 as a potential target in breast cancer and suggest combination with taxanes as a viable strategy to increase anti-tumor efficacy.