Cross-species genetic screens to identify kinase targets for APP reduction in Alzheimer's disease

Abstract

An early hallmark of Alzheimer's disease is the accumulation of amyloid-β (Aβ), inspiring numerous therapeutic strategies targeting this peptide. An alternative approach is to destabilize the amyloid beta precursor protein (APP) from which Aβ is derived. We interrogated innate pathways governing APP stability using a siRNA screen for modifiers whose own reduction diminished APP in human cell lines and transgenic Drosophila. As proof of principle, we validated PKCβ-a known modifier identified by the screen-in an APP transgenic mouse model. PKCβ was genetically targeted using a novel adeno-associated virus shuttle vector to deliver microRNA-adapted shRNA via intracranial injection. In vivo reduction of PKCβ initially diminished APP and delayed plaque formation. Despite persistent PKCβ suppression, the effect on APP and amyloid diminished over time. Our study advances this approach for mining druggable modifiers of disease-associated proteins, while cautioning that prolonged in vivo validation may be needed to reveal emergent limitations on efficacy.

Identification of risk genes for Alzheimer’s disease by gene embedding

“…Therefore, to optimally validate the GeneEMBED candidates in the AD context in vivo , we resorted to Drosophila AD models, which capture important core AD traits, including age dependence and protein accumulation. 89 This approach is supported by our previous Drosophila work in the context of AD and other neurodegeneration disorders, where therapeutic targets identified in Drosophila have gone on to be validated in mouse or induced pluripotent stem cell (iPSC)-derived neuronal models. 48 , 49 , 89 – 94 …”

“… 89 This approach is supported by our previous Drosophila work in the context of AD and other neurodegeneration disorders, where therapeutic targets identified in Drosophila have gone on to be validated in mouse or induced pluripotent stem cell (iPSC)-derived neuronal models. 48 , 49 , 89 – 94 …”

Temporal and spatially controlled APP transgene expression using Cre-dependent alleles

“…The work was framed around tetO-APP line 102 expressing the Swedish and Indiana mutations ( Jankowsky et al, 2005 ). In past work, this tetO-APP line had been bred directly with the very strong CaMKIIα-tTA driver line to elicit amyloid plaques within 1-2 months of birth ( Huichalaf et al, 2019 ; Rodgers et al, 2012 ). Here we wanted to control APP expression with Cre and thus relied on a Cre-to-tTA converter line to translate Cre activity into tTA expression.…”

“…The most common promoters for APP models turn on embryonically and remain active throughout life ( Jankowsky and Zheng, 2017 ). Amyloid formation in humans usually begins after 50 years of age, yet our most aggressive mouse models develop plaques before or soon after they can breed ( Chishti et al, 2001 ; Huichalaf et al, 2019 ; Oakley et al, 2006 ; Rodgers et al, 2012 ). Having the means to delay transgene onset might improve our modeling of this disease.…”

Acute silencing uncovers multiple forms of activity-dependent neuronal survival in the mature entorhinal cortex

“…Intracerebral viral injection was performed as previously described (Kim et al, 2014;Kim et al, 2016;Huichalaf et al, 2019). Neonatal Nop-tTA animals were stereotaxically injected at postnatal day 0 with 1 ul/hemisphere of AAV8 at a rate1 ul/min (5.4 x 10 9 gc/ul of TRE3G-hM4Di-T2a-YFP), targeting the lateral ventricles bilaterally: AP -1.0; ML +/-1.35; DV -1.7.…”