Persistent repression of tau in the brain using engineered zinc finger protein transcription factors

You'll have to ask your doctor if tau repression is more important than plaque removal to stop dementia.  And since this is in mice your doctor and hospital will need to ensure human testing. Or you could let them just be incompetent as usual and do nothing, but this impacts you and you might want a solution.

Persistent repression of tau in the brain using engineered zinc finger protein transcription factors

1. View ORCID ProfileSusanne Wegmann¹,²,*,†,
2. View ORCID ProfileSarah L. DeVos¹,*,
3. View ORCID ProfileBryan Zeitler³,*,†,
4. View ORCID ProfileKimberly Marlen³,
5. View ORCID ProfileRachel E. Bennett¹,
6. View ORCID ProfileMarta Perez-Rando¹,
7. View ORCID ProfileDanny MacKenzie¹,
8. Qi Yu³,
9. Caitlin Commins¹,
10. Riley N. Bannon¹,‡,
11. View ORCID ProfileBianca T. Corjuc¹,
12. View ORCID ProfileAlison Chase¹,
13. View ORCID ProfileLisa Diez²,
14. Hoang-Oanh B. Nguyen³,
15. View ORCID ProfileSarah Hinkley³,
16. View ORCID ProfileLei Zhang³,
17. Alicia Goodwin³,
18. Annemarie Ledeboer³,
19. View ORCID ProfileStephen Lam³,
20. Irina Ankoudinova³,
21. Hung Tran³,
22. View ORCID ProfileNicholas Scarlott³,
23. View ORCID ProfileRainier Amora³,
24. View ORCID ProfileRichard Surosky³,
25. Jeffrey C. Miller³,
26. View ORCID ProfileAshley B. Robbins¹,
27. Edward J. Rebar³,§,
28. View ORCID ProfileFyodor D. Urnov³,
29. View ORCID ProfileMichael C. Holmes³,
30. Amy M. Pooler³,
31. Brigit Riley³,
32. H. Steve Zhang³,|| and
33. View ORCID ProfileBradley T. Hyman¹,†

See all authors and affiliations

Science Advances  19 Mar 2021:
Vol. 7, no. 12, eabe1611
DOI: 10.1126/sciadv.abe1611

• Article
• Figures & Data
• Info & Metrics
• eLetters
• PDF

Abstract

Neuronal tau reduction confers resilience against β-amyloid and tau-related neurotoxicity in vitro and in vivo. Here, we introduce a novel translational approach to lower expression of the tau gene MAPT at the transcriptional level using gene-silencing zinc finger protein transcription factors (ZFP-TFs). Following a single administration of adeno-associated virus (AAV), either locally into the hippocampus or intravenously to enable whole-brain transduction, we selectively reduced tau messenger RNA and protein by 50 to 80% out to 11 months, the longest time point studied. Sustained tau lowering was achieved without detectable off-target effects, overt histopathological changes, or molecular alterations. Tau reduction with AAV ZFP-TFs was able to rescue neuronal damage around amyloid plaques in a mouse model of Alzheimer’s disease (APP/PS1 line). The highly specific, durable, and controlled knockdown of endogenous tau makes AAV-delivered ZFP-TFs a promising approach for the treatment of tau-related human brain diseases.

INTRODUCTION

The microtubule-binding protein tau is a key player in Alzheimer’s disease (AD) and frontotemporal dementia. The accumulation and aggregation of tau in the brain correlate with synaptic loss, neuronal loss, and cognitive decline (1–4). In patients with frontotemporal dementia, mutations in the tau gene, MAPT, lead to tau aggregation and cause widespread neurodegeneration (5), emphasizing the neurotoxic role of tau in these diseases. In addition to the neurotoxicity exerted by aggregated tau, soluble oligomeric forms of tau appear to be especially synaptotoxic (6). Mice engineered to lack expression of MAPT have been shown to be protected against β-amyloid (Aβ)–induced synaptotoxicity (7), as well as against stress-induced (8) and seizure-induced (9, 10) neuronal damage, and against learning and memory deficits resulting from traumatic brain injury (11). Moreover, reducing transgenic tau expression, even after tau has accumulated in mouse models of tauopathy, reverses the pathological effects of tau (12, 13). These findings support the idea that the reduction of tau protein could be used as a therapeutic approach in AD or other tauopathies.

Translation of the neuroprotective effect of tau repression into a therapeutic approach for neurodegenerative diseases requires a treatment that reduces endogenous tau in the adult brain. Previously, in vivo tau knockdown has been achieved through administration of antisense oligonucleotides (ASOs) that bind tau mRNA and prevent its translation (13, 14) or by immunotherapeutic intravenous injections of anti-tau antibodies (15–18). Although both approaches may facilitate tau protein reduction in the brain, they require chronic administrations to the patient and have limited ability to provide widespread knockdown.

Since recombinant adeno-associated viruses (rAAVs) can be used as viral vectors to efficiently transduce cells in the adult central nervous system (CNS) (19), we created a way to generate efficient, specific, and long-lasting down-regulation of the expression of endogenous tau by using a single viral administration: AAVs encoding engineered zinc finger protein (ZFP) arrays that precisely target a short region of the genomic mouse MAPT sequence and, through fusion to the KRAB repression domain of the human KOX1 transcription factor (TF), down-regulate MAPT gene expression. Using different AAV serotypes, we were able to reduce tau locally in the hippocampus—a brain region that is specifically affected by tau pathology in neurodegenerative diseases—through intracranial injections of AAV9 (adeno-associated virus serotype 9) or brain-wide through intravenous delivery of blood-brain barrier–crossing AAV-PHP.B (20). In both cases, a single AAV administration was sufficient to repress tau mRNA and all isoforms of the protein by 50 to 80% in the brain and for as long as we carried out the study—nearly 1 year—following the treatment.

Furthermore, we performed proof-of-principle experiments for the use of tau-targeted ZFP-TFs to treat neurodegeneration in a mouse model in vivo: The repression of endogenous tau appeared to protect neurons from toxicity in mice with AD-like Aβ pathology (APP/PS1 mice). Tau repression by ZFP-TFs reduced amyloid plaque–associated neuritic dystrophies, which are a tau-dependent pathological hallmark in these mice (21).

In summary, we present a novel translational approach to reduce tau in the adult brain, in which a single AAV ZFP-TF administration can produce potent, specific, and well-tolerated knockdown of endogenous neuronal tau. Our findings support the continued development of ZFP-TFs as a therapeutic platform for the treatment of tau protein–related disorders in the human brain.

RESULTS

Tau-targeted ZFP-TFs reduce endogenous tau expression in cultured primary mouse neurons

Tau reduction in the brain has been shown to be protective against pathological changes in different rodent models of neurodegeneration (7, 10, 22). To reduce all isoforms of tau in the adult mouse brain, we designed ZFPs—based on the backbone of human Zif268/EGR1—that target sequences within 500 base pairs of the transcription start site (TSS) of the mouse tau gene MAPT. The ZFPs efficiently binding the genomic MAPT sequence were then fused to a KRAB repression domain (TF). In each such ZFP-TF construct, the ZFP mediates the site-specific binding to the DNA, and the KRAB domain represses the endogenous expression of MAPT (Fig. 1A). When screening a panel of ~50 ZFP-TFs (out of 185 initial designs; fig. S1) for their dose-response activities on tau mRNA reduction in mouse neuroblastoma (N2a) cells, we identified candidates that robustly repressed tau mRNA and evaluated their off-target impact on global gene expression by RNA microarray. One candidate, ZFP-TF.89, targeting intron-1 of mouse MAPT (Fig. 1B), showed no detectable off-target transcriptome changes in N2a cells (fig. S1B and tables S1 and S2); we selected ZFP-TF.89 for further studies.

More graphs at link.