טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentYogev-Falach Merav
SubjectAmyloid Precursor Protein Processing and Neuroprotection
by the Anti-Alzheimer's Drug, Ladostigil (TV3326)
DepartmentDepartment of Medicine
Supervisor Professor Emeritus Moussa Youdim


Abstract

Two novel cholinesterase inhibitors (ChEIs), ladostigil (TV3326) and its S-isomer TV3279 [(N-propargyl-(3R) and (3S) aminoindan-5-yl)-ethyl methyl carbamate], were derived from rasagiline for the treatment of Alzheimer's disease (AD). Ladostigil, also inhibits selectively brain monoamine oxidase (MAO)-A and -B, whereas TV3279, lacks MAO inhibitory activity.

The primary aim of this work was to investigate the possibility that ladostigil and TV3279 may modulate the proteolytic processing of the amyloid precursor protein (APP). Here, we have shown that both isomers significantly stimulated the release of soluble APPa  (sAPPa) from both PC-12 and SH-SY5Y cells lines. Using several signal transduction inhibitors, we identified the involvement of protein kinase C (PKC) - and mitogen-activated protein kinase.

In vivo studies, demonstrated that administration of 150 mmoles/kg/14 days ladostigil or TV3279 to male C57/BL mice significantly decreased hippocampal APP levels. Furthermore, both drug up-regulated pPKC(pan) levels and the expression of a and e PKC isozymes in the mice hippocampus, indicating that the mechanism by which these drugs affect APP processing, in vivo,  may be related to PKC-associated signaling.  Similar to ladostigil, administration of 0.1 mg/kg/14 days rasagiline to mice significantly decreased hippocampal APP levels. Additionally, rasagiline up-regulated pPKC(pan) levels and the expression of a and e PKC isozymes. Moreover, similar to ladostigil, both rasagiline and N-propargylamine significantly increased the secretion of sAPPa via PKC and MAPK pathways, in vitro. These studies suggest the crucial role of the propargyl moiety in the effects of ladostigil, TV3279 and rasagiline.

We further assessed the neuroprotective activity of lodostigil, using high density apoptotic model in SK-N-SH cells.  Ladostigil significantly decreased apoptosis via inhibition of caspase-3 activation through a mechanism related to regulation of the Bcl-2 family proteins, resulting in reduced levels of Bad and Bax and induced levels of Bcl-2. Furthermore, ladostigil markedly decreased apoptotic-induced levels of holo-APP. Similar to ladostigil, its S-isomer, TV3279, exerted similar neuroprotective properties and APP processing, further suggesting that the mode of action is independent of MAO inhibition.  We also show that rasagiline or N-propargylamine almost completely inhibited apoptotic-induced caspase-3 cleavage and activation.  These results provide clear evidence that the neuroprotection effect of ladostigil is associated with the propargyl moiety in the compound activity on the mitochondria cell survival proteins.

Taken together, the neuroprotective effects of ladostigil and TV3279, together with their ability to induce the non-amyloidogenic sAPPa secretion via MAPK and PKC pathways, could clearly be of clinical importance for the treatment of AD.