DCP-LA , a New Strategy for Alzheimer ’ s Disease Therapy

Alzheimer’s disease (AD) is characterized by extensive deposition of amyloid β (Aβ) and formation of neurofibrillary tangles (NFTs) consisting of hyperphosphorylated Tau. So far, a variety of AD drugs targeting Aβ have been developed, but ended in failure. A recent focus on AD therapy, therefore, is development of Tau-targeted drugs. Aβ activates glycogen synthase kinase-3β (GSK-3β), that plays a central role in Tau phosphorylation, responsible for NFT formation. The linoleic acid derivative DCP-LA has been developed as a promising drug for AD therapy. DCP-LA serves as a selective activator of PKCε and a potent inhibitor of protein tyrosine phosphatase 1B (PTP1B). DCP-LA restrains Tau phosphorylation efficiently due to PKCε-mediated direct inactivation of GSK-3β, to PKCε/Akt-mediated inactivation of GSK-3β, and to receptor tyrosine kinase/insulin receptor substrate 1/phosphoinositide 3-kinase/3phosphoinositide-dependent protein kinase 1/Akt-mediated inactivation of GSK-3β in association with PTP1B inhibition. Moreover, DCP-LA ameliorates spatial learning and memory impairment in 5xFAD transgenic mice, an animal model of AD. Consequently, combination of PKCε activation and PTP1B inhibition must be an innovative strategy for AD therapy.


Introduction
Accumulating evidence has pointed to the role of amyloid  (A), a main body of amyloid (senile) plaques, and Tau protein, a main body of neurofibrillary tangles (NFTs), in the pathogenesis of Alzheimer's disease (AD).Huge studies have been done for development of AD drug targeting A, but no expecting drug has been obtained.Recent target, therefore, has been turned to Tau.
Tau is abundantly expressed in neurons of the central nervous system and stabilizes microtubules by interacting with tubulin.Microtubules are the tracks for motor proteins bearing intracellular transport of vesicles, organelles and protein complexes 1,2 , and Tau modulates microtubule dynamics including axonal transport [3][4][5][6] .Tau is upregulated during neuronal development, to promote generation of cell processes and establish cell polarity 7 .

Interaction between Aβ and GSK-3β
GSK-3 is originally in the active form.When phosphorylated at Ser9, GSK-3 is inactivated, but when phosphorylated at Tyr216, GSK-3 activation is enhanced 21 .
Amyloid precursor protein (APP) intracellular domain (AICD), that is produced from -secretase-mediated APP cleavage, activates GSK-3 29 or enters the nucleus and activates gene transcription, increasing the GSK-3 mRNA and protein 30 .C-terminal fragments of APP stimulate GSK-3 activation, to increase Tau phosphorylation and induce apoptosis 31 .
insulin or insulin-like growth-factor 1(IGF1) binds to and activates the RTK insulin receptor involving GSK-3 inactivation.
Aging, inflammation, and stress activate GSK-3, which triggers Tau phosphorylation, responsible for mild cognitive impairment (MCI), a preliminary group of AD (Figure 5).A further activates GSK-3 and accelerates Tau phosphorylation, leading to progression into AD from MCI (Figure 5) 43,44 .Aggregation of hyperphosphorylated Tau causes tauopathies, a class of neurodegenerative diseases, that include frontotemporal dementia and parkinsonism linked to chromosome 17, progressive supranuclear palsy, Pick's disease, and corticobasal degeneration as well as AD.Agents that have the potential to suppress GSK-3 activation, thus, could become beneficial preventive and therapeutic drugs for AD.
DCP-LA induces a long-lasting facilitation of hippocampal synaptic transmission by enhancing presynaptic 7 ACh receptor responses to stimulate glutamate release under the control of PKC [72][73][74][75] .In addition, DCP-LA activates CaMKII due to inhibition of protein phosphatase 1 (PP1), to enhance postsynaptic AMPA receptor responses and facilitate hippocampal synaptic transmission 76 .
The facilitatory action of DCP-LA on hippocampal synaptic transmission accounts for improvement of A 1-40 -and mutant A-induced spatial learning deficits in rats 77,78 , scopolamine-induced spatial learning and memory disorders in rats 77 , spatial learning and memory deterioration in senescence accelerated mice 8 (SAMP8) 79,80 , and spatial learning and memory impairment in 5xFAD transgenic mice, an animal model of AD 32 .
PKC is classified into the conventional PKC isozymes , I, II, and , the novel PKC isozymes , , , and , the atypical PKC isozymes / and , and the PKC-like isozymes  and .All the PKCs have the phosphatidylserine (PS) binding site and are activated by diacylglycerol (DG).Much interestingly, DCP-LA is capable of selectively activating PKC in a Ca 2+ -and DG-independent manner 81 .DCP-LA binds to the PS binding/associating sites Arg50 and Ile89 in the C2-like domain of PKC, which are distinct from the DG binding site in the C1 domain, at the carboxyl-terminal end and the cyclopropane rings, respectively 82 .
Racemic DCP-LA contains possible 4 diastereomers such as ,-, ,-, ,-, and ,-DCP-LA (Figure 6C).To develop DCP-LA as a medical drug, each diastereomer was separated and each characteristic was examined.Of 4 diastereomers ,-DCP-LA activates PKC selectively and stimulates presynaptic release of glutamate, dopamine, and serotonin, with the highest potency 83 .Of great interest is that DCP-PA serves as not only a selective PKC activator but a potent inhibitor of protein tyrosine phosphatase 1B (PTP1B).DCP-LA inhibits PTP1B by its direct interaction 84 .
PKC activation or PTP1B inhibition, thus, has the potential to restrain Tau phosphorylation by inactivating GSK-3 each independently.
Cooperation of PKC activation and PTP1B inhibition could inactivate GSK-3 and restrain Tau phosphorylation more efficiently than each solitary treatment 32 .In experiments using PC-12 cells, PKC overexpression and PTP1B deficiency activate Akt and inactivate GSK-3 synergistically 32 .A 1-42 activates GSK-3 by reducing Ser9 phosphorylation and increases Tau phosphorylation at Ser202/Thr205 and Ser396, and the effects of A 1-42 are clearly neutralized by DCP-LA 32 .
5xFAD mice are widely used as an animal model of AD. 5xFAD mice are APP/presenilin 1 (PS1) double transgenic mice that coexpress five familial forms of AD mutations such as the Swedish/London/Florida mutations and the M146L/L286V mutations 85 .The A 1-42 levels in the 5xFAD mouse brain increase in an age-dependent manner and spatial memory deficits are induced from 4-5 months of age 85 .The significantly higher levels of GSK-3-Ser9 phosphorylation is also found in the hippocampus of 5xFAD mice from 4-5 months of age as compared with the levels for wild-type control mice, indicating that the GSK-3 activity is enhanced in 5xFAD mice, possibly in association with A 1-42 increase 86 .Moreover, a greater deal of Tau-Ser396 phosphorylation, responsible for PHF formation, is found in the hippocampus of 5xFAD mice 86 .DCP-LA suppresses GSK-3 activation and reduces Tau-Ser396 phosphorylation in the hippocampus of 5xFAD mice to an extent similar to that for wild-type control mice 32 .DCP-LA, thus, enables efficient suppression of Tau-Ser396 hyperphosphorylation by activating PKC and inhibiting PTP1B simultaneously.DCP-LA ameliorates spatial learning and memory decline in 5xFAD mice, that occurs in parallel with GSK- activation and an increase in Tau phosphorylation, but such effect is not obtained with galanthamine, that is clinically used for treatment of mild to moderate AD 32 .In addition, DCP-LA improves A 1-40 -and mutant Ainduced spatial learning deficits in rats 77,78 , scopolamineinduced spatial learning and memory disorders in rats 77 , spatial learning and memory deterioration in senescence accelerated mice 79,80 .DCP-LA-induced improvement of cognitive decline is not due to only inhibition of GSK- and restraint of Tau phosphorylation.Facilitation of synaptic transmission in alive neurons would be required for improvement of cognitive decline.DCP-LA has the potential to facilitate hippocampal synaptic transmission by enhancing presynaptic 7 ACh receptor responses under the control of PKC [72][73][74][75] and postsynaptic AMPA receptor responses under the control of CaMKII in association with PP1 inhibition 76 .This action of DCP-LA is also a strong advantage as an AD therapeutic drug as compared with Tautargeted drugs including GSK- inhibitors.Tau-targeted drugs proposed possess no direct facilitatory action on synaptic transmission, and therefore, early improvement of cognitive decline would not be expected by those drugs.
A beneficial effect on 5xFAD mice is obtained with oral administration of DCP-LA at a dose of 1 mg/kg body weight, corresponding to ~3 M.This dose, in the light of the fact that the optimal concentration of DCP-LA in the in vitro experiments is 100 nM, seems to be appropriate and possible for clinical use.Overall, DCP-LA may shed a beam of hope on AD prevention and treatment.

Conclusion
Tau-targeted drugs for AD therapy under development include i) Hsp90 inhibitors, ii) inhibitors of A-induced Tau phosphorylation, iii) Tau aggregation inhibitors, iv) O-GlcNAcase inhibitors, v) GSK-3 inhibitors, vi) mTOR inhibitors, vii) inhibitors of Tau fibrillization, and viii) microtubule stabilizing agents.The mechanism underlying the inhibitory effect of DCP-LA on Tau phosphorylation is distinct from that for any drugs provided until now.DCP-LA restrains Tau phosphorylation efficiently due to PKCmediated direct inactivation of GSK-3, to PKC/Aktmediated inactivation of GSK-3, and to RTK/IRS-1/PI3K/ PDK1/Akt-mediated inactivation of GSK-3 in association with PTP1B inhibition.Consequently, combination of PKC

Figure 5 :
Figure 5: GSK-3 is a key factor for MCI and AD.Aging, inflammation, and stress activate GSK-3 and phosphorylate Tau, causing MCI.A enhances GSK-3 activation and accelerates Tau phosphorylation, leading to progression into AD from MCI.