Neuropharmacology

An In Vitro CNS Model to Test Compounds Counteracting Tau Pathology

The role of hyperphosphorylation of the microtubule-associated protein tau in pathological processes of several neurodegenerative diseases is becoming better understood. Consequently, development of new compounds capable of preventing tau hyperphosphorylation is an increasingly hot topic. For this reason reliable in vitroand in vivo models that reflect tau hyperphosphorylation in human diseases are needed.

To generate a new in vitro test system, the human neuroblastoma cell line SH-SY5Y was stably transfected with a pcDNA3-TMHT441 (V337M/R406W) construct. In these cells, tau expression is regulated by a CMV promoter. Over-expression of the TMHT441 construct leads to hyperphosphorylation of tau at different sites, such as Thr181, Ser202, Thr231/Ser235 and Ser396/Ser404, and is modifiable by kinase inhibitors, such as SP600125 and AR-A014418 (Löffler et al. 2012, J Mol Neurosci. PMID:22351109).

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Figure 1: Hyperphosphorylation of SH-SY5Y-TMHT441 cells as modified by different kinases. Total tau and ptau (Tau-5) expression levels in SH-SY5Y-TMHT441 cells compared with untransfected cells (wt) and TMHT mouse brain tissue at different residues (CTX and HC) (A);

Inhibition of pThr181 (B); and pThr231 (C) by kinase inhibitor SP60012; and inhibition of pSer396 by kinase inhibitor AR-A014418 (D). Inhibition of phosphorylation is shown relative to the levels of vehicle treated cells (B-D).

This in vitro cell system detects changes in the phosphorylation pattern of tau using several methods including immunosorbent assays and mass spectrometry. These changes can be reliably modulated by, for example, kinase inhibitors specifically targeting the protein kinases JNK, GSK-3, CDK1/5 and CK1. Therefore these cells represent an efficient tool for screenings and mechanistic studies. Other indication-specific cells can also be set up to facilitate lead finding for other diseases.

TMHT mice overexpressing TAU441 with the same mutations as the presented cell line are ready to proceed to an in vivo POC study (Flunkert et al. 2012, Neurodegener Dis. PMID: 22797329).

Authors:

QPS Austria
Birgit Hutter-Paier, PhD; Director Preclinical Department; 
birgit.hutter-paier@qps.com
 

QPS, LLC
 Walter Bee, PhD; VP & Head of Global Safety Assessment and Regulatory Affairs; walter.bee@qps.com

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