. Molecular basis for mid-region amyloid-β capture by leading Alzheimer's disease immunotherapies. Sci Rep. 2015 Apr 16;5:9649. PubMed.


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  1. Crespi and colleagues performed X-ray crystallography studies on two central domain-targeting anti-Aβ antibodies in order to identify structural interactions of the antibodies with the Aβ peptide (Crespi et al., 2015). Based upon the patent literature, the group developed their own “bio-similar” versions of solanezumab and crenezumab and subsequently investigated X-ray structural relationships in the presence of the Aβ fragment comprising amino acids 12 to 28. Their X-ray structural studies demonstrated that the antibody they produced corresponding to solanezumab bound to an Aβ epitope comprising amino acids 16 to 26, a finding that is consistent with our own internal epitope-mapping studies. Also, our epitope-mapping data indicates that the KLVFF domain (amino acids 16 to 20) by itself is not capable of binding to solanezumab by Biacore analyses. Of additional interest was the group’s finding that the Aβ peptide bound by the antibody had a central domain helical element that is quite different from the β-sheet structure found in oligomerized forms of the peptide. Our own studies have demonstrated the selectivity of solanezumab for soluble monomeric forms of the Aβ peptide (DeMattos et al., 2001; Racke et al., 2005). Our viewpoints diverge when the authors speculate in the discussion that the current structural work may explain their previously published observation of potential cross-reactivity of solanezumab and crenezumab to a number of proteins (Watt et al., 2014).

    We believe it is important to highlight that there were a number of technical issues with the author's earlier manuscript (Watt et al., 2014), especially as it pertains to the potential identification of cross-reacting proteins. These perspectives were highlighted in a letter to the editor by Lilly colleagues and several academic thought leaders (Siemers et al., 2014). To truly characterize a protein signature as “cross-reacting” from immunoprecipitation (IP) pull-down followed by MS/MS experiments on plasma, a matrix that has an immensely high protein concentration of 35 to 50 g/L, multiple rigorous controls and orthogonal biochemical approaches are necessary (binding assays, Biacore affinity measures, occupancy measures, and selectivity assays). Without rigorous orthogonal approaches, it is impossible to determine whether the signature was specific or merely representative of normal non-specific binding that would be expected for any globular protein. For example, would the authors believe the binding should be described as “cross-reacting” if they were to discover that it only represented a tiny fraction of the plasma pool (i.e. < 0.1 percent)? The specific binding of solanezumab results in an approximately 1000-fold increase in plasma Aβ (DeMattos et al., 2001). Indeed, our laboratory has recently analyzed non-specific plasma protein binding to other purified antibodies and we learned that hundreds, if not thousands of proteins can be detected in the IP-pull down experiments. Importantly, we analyzed these proteins by parallel approaches, such as SDS-PAGE followed by silver stain, where we determined that these non-specific protein interactions amounted to extremely low mass levels as compared to their concentration in plasma. Also, Crespi and colleagues conclude that the KLVFF homology is underlying the potential cross-reactivity (Crespi et al., 2015); however, our own epitope-mapping experiments in conjunction with Biacore analyses show that the KLVFF sequence is not capable of binding to solanezumab by itself.

    Furthermore, in the publications by Watt et al. and Crespi et al., the authors speculate that binding to the cross-reacting plasma proteins might result in a lack of target engagement of the central domain antibodies to plasma Aβ, and that if cognitive benefit is observed in current clinical studies, the effect could actually be due to unknown pharmacology linked to the cross-reacting proteins. To address the question of Aβ target engagement, in our subsequent letter to the editor we referenced an extensive literature from over two decades of work from multiple groups using multiple orthogonal biochemical approaches (multiple ELISA formats including multiple clinically validated assays, Western blotting, RIAs, acid urea gels, and MALDI mass spec) that demonstrate that the central domain anti-Aβ antibodies engage soluble Aβ in plasma and CSF  (DeMattos et al., 2001; Racke et al., 2005; Lachno et al., 2013; Siemers et al., 2010; DeMattos et al., 2002; DeMattos et al., 2002; Dodart et al., 2002; Yamada et al., 2009; Seubert et al., 2008; Bard et al., 2003; Seubert et al., 1992; Adolfsson et al., 2012; Doody et al., 2014; Farlow et al., 2012). These orthogonal approaches are more sensitive and quantitative than the SELDI methods employed by this group. Regarding the suggestion that binding to cross-reacting proteins leads to novel unknown pharmacology, we believe this hypothesis is premature, as their MS/MS experiments to explore this question did not adequately control for normal non-specific protein binding. Prior to this type of assertion, it would be prudent to perform the orthogonal biochemical approaches listed above. Moreover this hypothesis is inconsistent with the historical data for solanezumab referenced above.

    The clinical experience with solanezumab to date is consistent with a lack of meaningful binding to other proteins. Over 130,000 infusions of solanezumab have been administered to nearly 4,000 patients with AD, for durations in some cases over five years. The Phase 1, 2, and 3 clinical studies have documented robust target engagement (plasma and CSF) with good tolerability (Siemers et al., 2010; Doody et al., 2014; Farlow et al., 2012). The EXPEDITION 3 trial is now fully enrolled and the last patient visit is anticipated to occur around October 2016. If the EXPEDITION 3 trial confirms the slowing of cognitive and functional decline in mild AD that appeared to be present in EXPEDITION and EXPEDITION 2 pooled data (Carlson  et al., 2013), and also confirms what appears thus far to be good tolerability, these findings will support the amyloid cascade hypothesis and the targeting of soluble Aβ as one therapeutic avenue for the treatment of Alzheimer’s disease. 

    Eli Lilly's Michael Hutton, chief science officer, neurodegeneration, and Jirong Lu, senior research fellow, contributed to this comment.



    . Molecular basis for mid-region amyloid-β capture by leading Alzheimer's disease immunotherapies. Sci Rep. 2015 Apr 16;5:9649. PubMed.

    . Peripheral anti-A beta antibody alters CNS and plasma A beta clearance and decreases brain A beta burden in a mouse model of Alzheimer's disease. Proc Natl Acad Sci U S A. 2001 Jul 17;98(15):8850-5. Epub 2001 Jul 3 PubMed.

    . Exacerbation of cerebral amyloid angiopathy-associated microhemorrhage in amyloid precursor protein transgenic mice by immunotherapy is dependent on antibody recognition of deposited forms of amyloid beta. J Neurosci. 2005 Jan 19;25(3):629-36. PubMed.

    . Do current therapeutic anti-Aβ antibodies for Alzheimer's disease engage the target?. Acta Neuropathol. 2014 Jun;127(6):803-10. Epub 2014 May 7 PubMed.

    . Anti-Aβ antibody target engagement: commentary regarding Watt et al. Acta Neuropathol 127:803-810 (2014). Acta Neuropathol. 2014 Oct;128(4):609-10. Epub 2014 Aug 14 PubMed.

    . Validation of Assays for Measurement of Amyloid-β Peptides in Cerebrospinal Fluid and Plasma Specimens from Patients with Alzheimer's Disease Treated with Solanezumab. J Alzheimers Dis. 2013 Jan 1;34(4):897-910. PubMed.

    . Safety and changes in plasma and cerebrospinal fluid amyloid beta after a single administration of an amyloid beta monoclonal antibody in subjects with Alzheimer disease. Clin Neuropharmacol. 2010 Mar-Apr;33(2):67-73. PubMed.

    . Plaque-associated disruption of CSF and plasma amyloid-beta (Abeta) equilibrium in a mouse model of Alzheimer's disease. J Neurochem. 2002 Apr;81(2):229-36. PubMed.

    . Clusterin promotes amyloid plaque formation and is critical for neuritic toxicity in a mouse model of Alzheimer's disease. Proc Natl Acad Sci U S A. 2002 Aug 6;99(16):10843-8. PubMed.

    . Immunization reverses memory deficits without reducing brain Abeta burden in Alzheimer's disease model. Nat Neurosci. 2002 May;5(5):452-7. PubMed.

    . Abeta immunotherapy: intracerebral sequestration of Abeta by an anti-Abeta monoclonal antibody 266 with high affinity to soluble Abeta. J Neurosci. 2009 Sep 9;29(36):11393-8. PubMed.

    . Antibody capture of soluble Abeta does not reduce cortical Abeta amyloidosis in the PDAPP mouse. Neurodegener Dis. 2008;5(2):65-71. PubMed.

    . Epitope and isotype specificities of antibodies to beta -amyloid peptide for protection against Alzheimer's disease-like neuropathology. Proc Natl Acad Sci U S A. 2003 Feb 18;100(4):2023-8. Epub 2003 Feb 3 PubMed.

    . Peripherally administered antibodies against amyloid beta-peptide enter the central nervous system and reduce pathology in a mouse model of Alzheimer disease. Nat Med. 2000 Aug;6(8):916-9. PubMed.

    . Isolation and quantification of soluble Alzheimer's beta-peptide from biological fluids. Nature. 1992 Sep 24;359(6393):325-7. PubMed.

    . An effector-reduced anti-β-amyloid (Aβ) antibody with unique aβ binding properties promotes neuroprotection and glial engulfment of Aβ. J Neurosci. 2012 Jul 11;32(28):9677-89. PubMed.

    . Phase 3 trials of solanezumab for mild-to-moderate Alzheimer's disease. N Engl J Med. 2014 Jan 23;370(4):311-21. PubMed.

    . Safety and biomarker effects of solanezumab in patients with Alzheimer's disease. Alzheimers Dement. 2012 Jul;8(4):261-71. PubMed.

    . Efficacy of solanezumab in patients with mild or moderate Alzheimer's disease: Pooled analyses findings from two phase III studies. Alzheimer’s Dement. 2013 Jul;9 (4 Suppl):P139.

    View all comments by Eric Siemers
  2. Besides specific interaction details of the antibody with the amyloid peptide, one also has to consider the larger environment of Aβ synthesis, breakdown, forward and backward aggregation parameters, antibody-mediated microglia-dependent clearance, and impact on cognitive functioning through interaction with glutamate (Wang et al., 2013) and other neurotransmitter system such as the α7 nAchR (Lee et al., 2014). 

    Based on human SILK data, Aβ1-40 is synthesized faster and cleared more slowly than Aβ1-42 (Potter et al., 2013). Together with biochemical observations on the aggregation rates of these two amyloid peptides (Garai and Frieden, 2013), this results in a complex relationship between monomers, oligomers, and aggregated forms of both peptides. Assuming also that oligomers might be formed from breakdown of plaques, it is clear that many non-linear processes play a role and that the clinical outcome might depend upon the specific epitopes and their affinities of the antibodies.

    Recently we showed that the choice of the epitope for the antibody does matter, both in terms of biomarkers (CSF Aβ levels) and functional outcome (effect on ADAS-Cog). Our approach was a mechanism-based Quantitative Systems Pharmacology and humanized computer model, constrained by existing clinical data on amyloid modulation in AD patients based on integration of biological, biochemical and clinical domain expertise (Geerts 2014).




    . Increased in vivo amyloid-β42 production, exchange, and loss in presenilin mutation carriers. Sci Transl Med. 2013 Jun 12;5(189):189ra77. PubMed.

    . Quantitative analysis of the time course of Aβ oligomerization and subsequent growth steps using tetramethylrhodamine-labeled Aβ. Proc Natl Acad Sci U S A. 2013 Feb 26;110(9):3321-6. PubMed.

    . Multiple effects of β-amyloid on single excitatory synaptic connections in the PFC. Front Cell Neurosci. 2013;7:129. Epub 2013 Sep 3 PubMed.

    . Picomolar Amyloid-β Peptides Enhance Spontaneous Astrocyte Calcium Transients. J Alzheimers Dis. 2014 Jan 1;38(1):49-62. PubMed.

    . Using in silico mechanistic disease modeling to address the effect of amyloid beta manipulation on cognitive clinical readouts. Alzheimers Dement. 2011 Jul; 7(4 Suppl),S774–5.

    View all comments by Hugo Geerts

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  1. Shape of a Hug: How the Embrace of a Therapeutic Aβ Antibody Really Matters