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Liao F, Hori Y, Hudry E, Bauer AQ, Jiang H, Mahan TE, Lefton KB, Zhang TJ, Dearborn JT, Kim J, Culver JP, Betensky R, Wozniak DF, Hyman BT, Holtzman DM. Anti-ApoE antibody given after plaque onset decreases Aβ accumulation and improves brain function in a mouse model of Aβ amyloidosis. J Neurosci. 2014 May 21;34(21):7281-92. PubMed.
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Indiana University School of Medicine
The report by Michaelson's lab provides further validation of the effects of the RXR agonist bexarotene in murine models related to AD. They have employed a mouse model in which the human ApoE genes are knocked into the murine locus and, unlike previous studies, only the murine Aβ species are linked to the deficits in these mice. Importantly, they find bexarotene-mediated improvement in memory and learning in two different behavioral tasks, consistent with outcomes reported by Cramer et al., Fitz et al., and Tesseur et al. They report a drug-induced reduction in Aβ42 levels, similar to those reported by us (Cramer et al., 2012).
Significantly, they have explored the effects of bexarotene on neurons and find that RXR activation reduces phosphorylated tau detected by the antibody AT8. Moreover, they argue that the drug has effects on the presynaptic marker VGlut1, elevating its abundance in the ApoE4 mice. There are a few unanswered questions. It is not clear why Aβ42 should be preferentially cleared and not Aβ40. They show a bexarotene-mediated reduction in AT8 immunoreactivity, but it is entirely unclear how this effect on tau is achieved. This work is noteworthy as it addresses the effects of bexarotene in a model that does not develop plaques, providing new evidence that plaques are functionally irrelevant.
The Holtzman paper is very intriguing and follows from their 2012 paper. The new work reinforces the view that ApoE contributes in significant ways to regulation of amyloid deposition and that manipulation of ApoE levels with immunotherapy, or direct antibody exposure, alters plaque dynamics. The data suggest that immunotherapy applied after disease onset may be of therapeutic utility—at least in mice. The best effects on plaque reduction were seen in the thalamus, not a typical area evaluated in AD, with smaller or no effects seen in the cortex and hippocampus, respectively. The paper contains a few odd findings, including an effect on motor endpoints. The analysis of functional connectivity is a high-tech flourish that augments the principal findings of the paper. This is a nicely performed study that takes advantage of the substantial technical resources available to the Holtzman and Hyman labs. The caliber of the work is excellent and provides direct support for ApoE-based therapeutics.
References:
Cramer PE, Cirrito JR, Wesson DW, Lee CY, Karlo JC, Zinn AE, Casali BT, Restivo JL, Goebel WD, James MJ, Brunden KR, Wilson DA, Landreth GE. ApoE-directed therapeutics rapidly clear β-amyloid and reverse deficits in AD mouse models. Science. 2012 Mar 23;335(6075):1503-6. Epub 2012 Feb 9 PubMed.
Fitz NF, Cronican AA, Lefterov I, Koldamova R. Comment on "ApoE-directed therapeutics rapidly clear β-amyloid and reverse deficits in AD mouse models". Science. 2013 May 24;340(6135):924-c. PubMed.
Tesseur I, Lo AC, Roberfroid A, Dietvorst S, Van Broeck B, Borgers M, Gijsen H, Moechars D, Mercken M, Kemp J, D'Hooge R, De Strooper B. Comment on "ApoE-directed therapeutics rapidly clear β-amyloid and reverse deficits in AD mouse models". Science. 2013 May 24;340(6135):924-e. PubMed.
View all comments by Gary LandrethMayo Clinic
Both these studies are addressing the ApoE problem by using different strategies to alter ApoE concentrations.
Liao and colleagues used a monoclonal mouse anti-ApoE antibody trying to sequester ApoE levels in plaque-bearing APP/PS1 mice at the age of 7 months. Using intraperitoneal injection of the HJ6.3 antibody for 21 weeks, the authors describe decreased Aβ plaque load, increased plasma Aβ levels, improved spatial learning, and decreased microglial activation in the cerebral cortex. These findings are very interesting given the continuous debate about whether to increase or decrease ApoE levels to combat its pathogenic effects. Interestingly, there was no effect on plasma ApoE levels upon immunization; there was, however, a slight reduction in ApoE from Triton-X soluble fractions from brain tissue lysates. The authors also did not observe any difference in plasma cholesterol. These latter results are surprising, as theoretically the administration site (periphery) would promote peripheral levels of ApoE as the first target upon antibody injection. It would be very interesting to figure out the titer distribution of this antibody. How much of it actually reaches the brain? The results from this study are very encouraging and if they hold true, it would mean that only a slight reduction of ApoE in a specific ApoE pool is needed to get beneficial effects.
In the second study, Boehm-Cagan and Michaelson have investigated the effect of the highly debated bexarotene on mRNA and protein levels of ApoE and the lipid transporters ABCA1 and ABCG1 in ApoE3 and ApoE4 targeted replacement mice. Contrary to earlier published in vitro data (Cramer et al., 2012) they describe no effect of bexarotene on mRNA and protein levels of ApoE. They do, however, describe an increase in both ABCA1 and ABCG1. Importantly, they found that the lipidation of ApoE4, as determined by immunoblotting, increased upon bexarotene treatment. The authors describe reversal of cognitive deficits in ApoE4 mice, and a reduction in Aβ accumulation and AT8 tau phosphorylation in both ApoE3 and ApoE4 mice.
The benefits described in both these studies appear to happen without any major influence on ApoE protein concentrations. This is intriguing. From our own studies on AD patients and controls, we know that CSF levels of total ApoE don’t differ between APOE genotypes. Similarly, the distribution of individual ApoE isoforms is about 50-50 in CSF from APOE3/4 carriers (Martinez-Morillo et al., 2014). Together these various findings suggest that not the levels of ApoE per se are important for AD pathology but rather qualitative characteristics—as suggested by Boehm-Cagan and Michaelson.
Last, in their discussion Boehm-Cagan and Michaelson mention that it’s important to note that bexarotene has peripheral effects on, for instance, lipid levels. This point is crucial, as the peripheral mechanisms are too often excluded from the discussion. For example, we recently found that despite the unaltered levels of the ApoE4 isoform in CSF from APOE4 carriers, the same individuals did exhibit a specific decrease in ApoE4 levels in plasma. In heterozygotes, the levels of the non-ApoE4 isoform were unaltered (Martinez-Morillo et al., 2014), thus demonstrating that the observed decrease in total plasma ApoE in APOE4 carriers is attributed to a specific decrease in the ApoE4 isoform.
Importantly, there are other described effects of rexinoids that are rarely mentioned. A Nature publication back in 1997 described antidiabetic effects of RXR ligands, which functioned as insulin sensitizers decreasing fasting glucose levels and reducing both hyperglycemia and hyperinsulinemia in mouse models of diabetes and obesity (Mukherjee et al., 1997). In regard to the documented effects on insulin on cognition, the anti-diabetic effects of bexarotene need to be looked into further.
References:
Cramer PE, Cirrito JR, Wesson DW, Lee CY, Karlo JC, Zinn AE, Casali BT, Restivo JL, Goebel WD, James MJ, Brunden KR, Wilson DA, Landreth GE. ApoE-directed therapeutics rapidly clear β-amyloid and reverse deficits in AD mouse models. Science. 2012 Mar 23;335(6075):1503-6. Epub 2012 Feb 9 PubMed.
Martínez-Morillo E, Hansson O, Atagi Y, Bu G, Minthon L, Diamandis EP, Nielsen HM. Total apolipoprotein E levels and specific isoform composition in cerebrospinal fluid and plasma from Alzheimer's disease patients and controls. Acta Neuropathol. 2014 May;127(5):633-43. Epub 2014 Mar 15 PubMed.
Mukherjee R, Davies PJ, Crombie DL, Bischoff ED, Cesario RM, Jow L, Hamann LG, Boehm MF, Mondon CE, Nadzan AM, Paterniti JR Jr, Heyman RA. Sensitization of diabetic and obese mice to insulin by retinoid X receptor agonists. Nature. 1997 Mar 27;386(6623):407-10. PubMed.
View all comments by Henrietta NielsenUniversity of Illinois, Chicago
University of Illinois
As APOE4 is basically the genetic risk factor for AD, it is heartening to see these two papers.
The paper by Holtzman and collaborators is a characterization that leaves few questions unanswered. That plaque burden decreases with systemic delivery of an ApoE antibody is very interesting. Coupled with the biochemical extraction that shows decreases in soluble Aβ and Aβ in the plasma, immunochistochemistry for microglial activation, extensive behavioral analysis, and evidence that lipoprotein biogenesis is not significantly disrupted by the removal of ApoE, the news here is excellent. The addition of the topical application to live mice and the subsequent reduction in plaques is very compelling and adds another dimension to the interpretation of the results. It will be interesting to see what effect the human APOE genotypes have on this antibody therapy.
An ongoing debate in the field is whether ApoE4 represents a toxic gain or loss of function, and how this distinction will impact the activity of APOE-directed AD therapeutics. Michaelson and collaborators address this issue through treating ApoE-targeted replacement mice, which express mouse Aβ and human ApoE3 or ApoE4, with the anti-cancer drug bexarotene. Bexarotene, through increasing ABCA1/G1 expression, reversed the lipidation deficiency associated with ApoE4, without any effect on total ApoE levels. Importantly, bexarotene lowered intraneuronal Aβ42 and hyperphosphorylated tau levels and reversed cognitive deficits only in ApoE4-TR mice. These data provide compelling evidence that increasing lipoprotein lipidation in the CNS may reverse the loss of function associated with ApoE4. To capitalize on these findings, further research on the effects of APOE, human Aβ and ApoE-lipidation on AD progression are sorely needed.
View all comments by Leon TaiCase Western Reserve University
There has been a growing realization that we need to expand AD drug targets beyond amyloid. Therefore, at first glance the news of ApoE as a potential therapeutic target would be welcomed. As other commentators have pointed out, the Holtzman paper presents solid data, based on state-of-the-art technology, to establish that ApoE immune therapy hits all the right milestones in APP/PS1 mice—decreased amyloid plaque load, improved spatial learning in Morris water maze etc.—and could be a potential treatment for human AD. However, upon some reflection it becomes less clear why this approach should prove effective in AD patients when the past efforts to lower Aβ plaque load by passive immunotherapy—which worked brilliantly in mice—failed in clinical trials.
Notwithstanding the beautiful science, the rationale for ApoE immunotherapy in humans remains weak because it is the E4 variant of ApoE and not ApoE protein itself that increases AD risk. Moreover, although Aβ is invoked, the precise mechanism by which ApoE4 increases AD risk remains unclear. Young adult ApoE4 carriers also exhibit altered connectivity of the medial temporal lobes (Dennis et al., 2010), increased seizure susceptibility (Briellmann et al., 2000), propensity for HIV dementia (Corder et al., 1998) and impaired recovery following traumatic brain injury (Alexander et al., 2007), all of which are independent of Aβ. The AD field must look beyond Aβ to find an effective disease modifying treatment.
On a side note, the authors showed that mice treated with anti-apoE antibody exhibited improved performance in a Morris water maze test (which is strongly dependent on hippocampal synaptic plasticity) without any decrease in hippocampal amyloid plaque load. Should we believe that hippocampal plaques do not matter?
References:
Dennis NA, Browndyke JN, Stokes J, Need A, Burke JR, Welsh-Bohmer KA, Cabeza R. Temporal lobe functional activity and connectivity in young adult APOE varepsilon4 carriers. Alzheimers Dement. 2010 Jul;6(4):303-11. PubMed.
Briellmann RS, Torn-Broers Y, Busuttil BE, Major BJ, Kalnins RM, Olsen M, Jackson GD, Frauman AG, Berkovic SF. APOE epsilon4 genotype is associated with an earlier onset of chronic temporal lobe epilepsy. Neurology. 2000 Aug 8;55(3):435-7. PubMed.
Corder EH, Robertson K, Lannfelt L, Bogdanovic N, Eggertsen G, Wilkins J, Hall C. HIV-infected subjects with the E4 allele for APOE have excess dementia and peripheral neuropathy. Nat Med. 1998 Oct;4(10):1182-4. PubMed.
Alexander S, Kerr ME, Kim Y, Kamboh MI, Beers SR, Conley YP. Apolipoprotein E4 allele presence and functional outcome after severe traumatic brain injury. J Neurotrauma. 2007 May;24(5):790-7. PubMed.
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