A presentation on May 7th 2015 at the Epigenomics Journal Club we had started. This one comprises the main papers looking at Alzheimer's Disease and the immuno-cell specific signatures shown in the mice models. A complementary study in patients found ANK1 as a clear EWAS association.

1. Epigenomics JC 150507
Gjoneska et al. 2015 Nature 518(365)
Lunnon et al. 2015 Nature Neurosci.17(9)
Álvaro Martínez Barrio, PhD
martinezbarrio.alvaro@gmail.com
linkedin.com/in/ambarrio
@ambarrio

3. Extended Figure 1

7. Extended Figure 1
Cyclin-dependent kinase 5,
regulatory subunit 1
(CDK5R1 -> p35)
cyclin-dependent kinase 5 (CDK5)

8. Extended Figure 1
Cyclin-dependent kinase 5,
regulatory subunit 1
(CDK5R1 -> p35)
cyclin-dependent kinase 5 (CDK5)
calpain
(p25)
+ relocalization from the cell periphery to nuclear and perinuclear regions

9. Extended Figure 1
Cyclin-dependent kinase 5,
regulatory subunit 1
(CDK5R1 -> p35)
cyclin-dependent kinase 5 (CDK5)
(p25)
+ deregulates CDK5 activity by prolonging its
activation and changing its cellular location.
+ p25 form accumulates in the brain neurons of
patients with AD.
+ accumulation correlates with an increase in
CDK5 kinase activity
+ may lead to aberrantly phosphorylated forms
of the microtubule-associated protein tau,
which contributes to AD.

10. Extended Figure 1
Cyclin-dependent kinase 5,
regulatory subunit 1
(CDK5R1 -> p35)
cyclin-dependent kinase 5 (CDK5)
(p25)

11. Extended Figure 1
+ Triple transgenic (Tg) mice over-expressing human p35,
Cdk5, and tau, despite significant increases in Cdk5
activity in these mice, do not display an increase in tau
phosphorylation (Van den Haute et al., 2001).
!
+ Generation of Inducible Tg Mice overexpressing human
p25 in the Postnatal Forebrain using the tetracycline-
controlled transactivator (tTA) system under the control
of the CamKII promoter, which drives high transgene
expression in the forebrain (Mayford et al., 1996).
!
+ In the presence of the tetracycline derivative
doxycycline, expression of the p25 transgene is
inhibited.
!
+ All mice are conceived and raised in the presence of
doxycycline beforeinduction of p25 to prevent any
potential developmental consequences from the
expression of p25.
5. Cruz, J. C., Tseng, H.-C., Goldman, J. A., Shih, H. & Tsai,
L.-H. Aberrant Cdk5 activation by p25 triggers pathological
events leading to neurodegeneration and neurofibrillary
tangles (NFTs). Neuron 40, 471–483 (2003).

12. Figure 1

13. Extended Figure 9 Extended Figure 2

14. Figure 1
2. Blalock, E. M., Buechel, H. M., Popovic, J., Geddes, J. W. &
Landfield, P. W. Microarray analyses of laser-captured
hippocampus reveal distinct gray and white matter signatures
associated with incipient Alzheimer’s disease. J. Chem.
Neuroanat. 42, 118–126 (2011).

15. Extended Figure 1 Extended Figure 3

16. Extended Figure 1 Extended Figure 3

17. Extended Figure 4

18. Extended Figure 5
Figure 1

19. Figure 1
Extended Figure 1

20. Figure 3
Figure 2

21. Figure 2

22. Extended Figure 6

23. 4. Lambert, J. C. et al. Meta-analysis of 74,046 individuals
identifies 11 new susceptibility loci for Alzheimer’s disease.
Nature Genet. 45, 1452–1458 (2013).
Figure 2
Extended Figure 7

24. Extended Figure 8

25. Figure 3
+ Among 20 genome-wide significant AD loci, 11 non-coding AD-associated SNP in LD. 5/11 localise w/i increased-level
enhancers (e.g. PICALM, BIN1) and three (INPP5D, CELF1/SPI1, PTK2B) by combining all AD cohorts.
!
+ Additional 22 weakly associated regions contains variants w/i increased-level enhancer orthologues, 17 lack protein-altering
variants in LD (e.g ABCA1, prologue of AD-associated ABCA7).

26. Figure 3
29. Krimbou, L. et al. Molecular interactions between apoE and ABCA1:
impact on apoE lipidation. J. Lipid Res. 45, 839–848 (2004).
Apolipoprotein E (ApoE)
+ Class of apolipoprotein found in the chylomicron and
Intermediate-density lipoprotein (IDLs) that is essential
for the normal catabolism of triglyceride-rich
lipoprotein constituents.
+ In peripheral tissues, ApoE is primarily produced by
the liver and macrophages, and mediates cholesterol
metabolism in an isoform-dependent manner.
+ In the CNS, ApoE is mainly produced by astrocytes, and
transports cholesterol to neurons via ApoE receptors,
which are members of the low density lipoprotein
receptor gene family.

28. + AD affects 26m people worldwide.
!
+ Accumulation of amyloid plaques > NFT by hyperphosphorylated tau >
gliosis > synaptic dysfunction > neuronal cell death
!
+ Progressive neurodegeneration across the cortex, with areas of early
neuropathology as the entorhinal cortex (EC) and others resistant to
neuronal damage like the cerebellum (CER).
!
+ Etiology of AD focused primarily on DNA sequence variation. Jonathan Mill
is an advocate of epigenome-wide association studies (EWAS).

29. • Study DESIGN: one discovery cohort (n=122) with
four brain regions (EC, STC, PFC, CER and blood pre-
mortem), 2 confirmation ones (n=144 and n=62).
!
• TOOL: Illumina 450K HumanMethylation array and
some targeted bisulfite-pyrosequencing.
!
• CATEGORIES: Braak staging (0-VI), a standarized
measure of neurofibrillary tangle burden
determined at autopsy.

31. Figure 1

32. Figure 1

33. Figure 2
Supplementary Figure 1

34. Figure 2

35. Figure 2
+ EWAS of AD pathology in 708 cortical samples
21. De Jager, P.L. et al. Alzheimer’s disease pathology is
associated with early alterations in brain DNA methylation at
ANK1, BIN1 and other loci. Nat. Neurosci. doi:10.1038/nn.
3786 (17 August 2014).
Supplementary Figure 2
+ Although not previously implicated in dementia, genetic variation in ANK1 is
associated with diabetic phenotypes, establishing links between T2D and
AD.

36. Supplementary Figure 3
Supplementary Figure 4 (N=36)

37. + Clinical biomarkers with diagnostic and prognostic utility during AD-development.
+ Pre-mortem whole-blood DNA from the discovery cohort (n=57).
+ Duration between blood sampling and mortality (avg = 4.15 ± 3.00 years).
+ Analyses restricted to identification of DMPs associated with clinical diagnosis of AD instead of Braak score.
+ Identify a number if AD-associated DMPs, many in the vicinity of relevant to AD genes (e.g. DAPK1, GAS1, NDUFS5).
+ However, the top-ranked DMPs in blood are distinct from those identified in the brain as they show no significant overlap with either
cortex or CER.
+ AD-associated DMPs in blood are unlikely related to the actual neurodegenerative process.
+ However, using previous blood-based transcriptomic data (ref. 38), they observe 18 of their top-ranked blood DMPs located in the vicinity
of known differentially expressed transcripts.

39. Figure 4

40. Gjoneska et al. Lunnon et al. 2015
• Contrasting changes in immune and
neuronal genes and regulatory regions
during AD-like degeneration
• Surprising depletion of neuronal
promoters and enhancers for a cognitive
disorder with well-established
connections to environmental and
experiential factors
• Model of interactions between genetically
driven immune cell dysregulation and
environmentally driven epigenetic
alteration in neural cells
• Power of the CK-p25 mouse model for the
study of AD human disease progression.
Molecular changes in both genes and
regulatory regions are highly conserved.
• Specific therapeutic targets for AD (e.g.
PU.1)
• Largest cross-tissue study of AD using matched DNA
from both affected and unaffected brain regions.
First sequential replication with 3 independent
study cohorts and 2 independent technologies.
• Cortex-specific hypermethylation across an ANK1
region in several independent cohorts suggestive of
the locus being relevant to AD-pathogenesis. Most
substantial in EC. Replication of the same DMR in
another large EWAS of AD (De Jager et al.).
• Most brain-identified DMPs, including ANK1, not
detected in blood. But many DMPs in blood located
in vicinity of genes found altered in patients with
MCI.
• Careful control of technical artefacts in EWAS with
Illumina 450K array.
• Power calculations from EWAS in study design. They
state to be powered to identify relatively small
(~5%) DNA methylation differences between
groups.
• Conventional methods for multiple-test correction
likely to be overly stringent and inappropriate given
non-independence of loci and lack of inter-
individual variation.