The significance of A and

The significance of Aβ and tau as therapeutic targets is not only derived from pathological evidence of postmortem brain but also from biochemical analysis of cultured human neurons. This is largely based on the induced pluripotent stem cell (iPSC) technology that has been established to study Aβ, tau, and GSK3β in AD [28]. When iPSC lines from two normal subjects, two SAD (sAD1 and sAD2), and two FAD patients carrying a duplicated copy of the APP gene were established for human neuronal differentiation, those from two duplicate APP gene carriers and patient sAD2 showed very high levels of Aβ40, phosphor-tau(Thr 231) and active GSK3β [28]. Importantly, levels of Aβ, pTau and active GSK3β can be reduced by β-secretase inhibitors, indicating a direct relationship between γ-secretase substrate C99 and GSK3β activation/Tau phosphorylation. Other studies have demonstrated that iPSC-derived neuronal cells exhibit reduced Aβ levels in the presence of BACE1 inhibitor or γ-secretase inhibitor/modulators [29,30]. Thus, involvement of Aβ and tau in AD pathogenesis can be modeled in human neuronal cells amendable for testing of therapies targeting either Aβ or phosphorylated tau [31].
Targeting γ-secretase with inhibitors: from the end to the beginning? Aβ targeted therapies are being actively pursued in preclinical and clinical studies for treatment of AD. These therapies are based on the amyloid cascade hypothesis, which postulates that Aβ peptides form neurotoxic species, trigger a pathological cascade and ultimately lead to neurodegeneration and dementia [32,33]. γ-Secretase, along with the β-secretase, have become the prime target for this purpose. γ-Secretase is composed of PS1, Presenilin Enhancer-2 (Pen-2), anterior pharynx defective-1 (Aph-1), and Nicastrin (Nct). PS1 carries two aspartate residues constituting the active site of γ-secretase [34], and Pen-2 is a small protein of 101 doxercalciferol with two TM domains [35,36]. Many studies have demonstrated that overexpression of all four components results in increased γ-secretase activity, both in mammalian cells [[37], [38], [39], [40], [41], [42]] and in yeast [43]. Purified PS1 and Pen-2 are sufficient to carry out γ-secretase cleavage of its substrates in vitro; PS1 itself has proteolytic activity [44,45], while Pen-2 promotes the conversion of PS1 from zymogen to the active protease [44,46]. A large number of potent γ-secretase inhibitors (GSIs) have been developed. Two GSIs, LY-450139 (Semagacestat) and BMS-708163 (Avagacestat), are among those tested in clinical trials. LY-450139 is known to block the cleavage of APP and Notch, another γ-secretase substrate [47], and subjects receiving LY-450139 presented with worsening of clinical measures of cognition and the ability to perform activities of daily living [48,49]. Because perturbed Notch signaling has been implicated in cancer formation, inhibition of Notch signaling by LY-450139 could be one of the culprits causing the undesired clinical outcomes [50]. More selective GSIs targeting APP over Notch were developed, like allosteric GSIs and BMS-708163. Allosteric GSIs (AGSI) displays specificity against Aβ42 production over Aβ40, Aβ38 and Notch1 cleavage [51]. These AGSIs bind to an allosteric site within γ-secretase rather than the APP substrate. Furthermore, AGSIs affect γ-secretase activity for both Aβ40 and Aβ38 production similarly and therefore lack the interconnected effect of the γ-secretase modulators (GSM) (see below) in which decreased Aβ42 resulted in increased Aβ38 generation [52]. Clearly, these AGSIs represent a class of inhibitors that are distinct from the Notch-sparing GSIs that have no selectivity for Aβ40 and Aβ42 [[53], [54], [55]]. BMS-708163 is a potent GSI that showed impressive γ-secretase inhibition with 50% inhibition concentrations (IC50) of 0.27 and 0.30 nM for Aβ42 and Aβ40, and 58 nM for Notch, respectively, thus representing a 193-fold selectivity for APP over Notch [56]. Pharmacokinetic (PK)-pharmacodynamic (PD) analysis of BMS-708163 in male dogs revealed plasma concentration of BMS-708163 of ∼0.5 μM at 3 h post-dosing (hpd) and brain concentration of ∼0.75 μM at 5 hpd, with a sustained decrease of brain Aβ40 by 50%. However, a separate study indicates that only 3–7 fold selectivity exists for APP over Notch [57], and clinical outcomes [58,59] seem to be discrepant with reported 193-fold selectivity for APP over Notch cleavage [56].