Lo Cascio et al JBC 295, 14807-25, 2020

 Modulating disease-relevant tau oligomeric strains by small molecules

Filipa Lo Cascio, Stephanie Garcia, Mauro Montalbano, Nicha Puangmalai, Salome McAllen, Andrea Pace, Antonio Palumbo Piccionello, Rakez Kayed

claim curcumin derivitives bind and change the conformation of toxic tau oligomers into larger, less toxic aggregates

something to look at with our compounds

disease relevant brain derived tau oligomers from AD, DLB, and PSP by IP with T22 antibody then amplified using full length 2N4R recombinant tau

primary cortical neurons exposed to 0.5uM BDTOs alone or BDTOs pretreated with 5uM compound for 24h, measure cyttoxicity by LDH - we could try this

Preparation of recombinant tau oligomers for cell assay studies:

300 μl of tau stock (0.3 mg) was added to 700 μl of 1× PBS and incubated for 1 h on an orbital shaker at room temperature. After shaking, the resulting TauO were purified by FPLC (Superdex 200 Increase 10/300 column, Amersham Biosciences).

primary cells used C57BL/6 mice (Jackson Laboratory, stock number 000664) exposed to brainderived oligomers 0.125 to 1µM +/- 5µM compound for 24h, then MTS or LDH viability assay

disagree with statement of different proteinase patterns of AD vs DLB vs PSP in Fig 1 E and F, looks more like difference in quantity rather than species. however, the band patterns in undigested (Fig1d) certainly look different

densitometry in Fig 2 does not seem to match with blots shown, some bands are distinctly more dense than the control and yet the quantification shows no significant difference. conversely, some bands are less than half the density of the control but the quantification shows only a 10% drop

fig 3 shows a change in band pattern of BDTO as a result incubation with compound. a decrease in higher molecular weight bands without a corresponding increase in monomers is supportive that the compound causes an increase in very large aggregates (>250kDa)

fig 4 drives me crazy due to the changing scales but again supports generation of large aggregates

fig 5 shows compound partial rescue of BDTO induced toxicity in primary neurons - by IF

fig 7 is intriguing subcellular fractionation after exposure of primary neurons to BDTO +/-preincubation with compound - a decrease in monomeric tau is found in all 3 compartments when incubated with oligomers. authors claim this is evidence of seeding activity. densitometry data again makes no sense, opposite results with cmpound in AD vs PSP, very confusing, but a very interesting concept

fig 8 is using biosensor seeding assay: transfect with BDTOs in Lipofectamine 2000 6µL Lipo, incubate 24 hours, analyze by IF