C224 thematic

Alzheimer’s: biomarkers redefine disease

Biological markers (biomarkers) for Alzheimer’s disease (AD) assess risk as well as the presence and progression of the disease.
AD biomarkers are derived from cerebrospinal fluid (CSF) and plasma.  Some biomarkers have been evaluated as part of regulatory guideline documents, often in Phase II drug development trials establishing safety and tolerance. Large-scale multi-centre trials are expected to validate biomarker candidates for use in Phase III studies.
Nevertheless, AD biomarkers face several hurdles which they must cross before attaining their full potential.

Challenges of timely or definitive diagnosis
The timely diagnosis of AD is difficult because its early symptoms are shared by a variety of disorders with similar neuropathological features. These include vascular dementia (VaD), frontotemporal lobe dementia (FTLD) and Lewy body dementia (LBD).  Failure to distinguish AD from the others can be a challenge since clinicians are likely to select different approaches to treatment. Still, given that AD-related molecular mechanisms precede symptoms, biomarkers have the potential to be used as early indicators and markers of pre-clinical pathological change.

One of the major, underlying challenges with AD is that definitive diagnosis is not possible so far in living patients. Making such a diagnosis requires clinical assessment of AD via biomarkers as well as post-mortem verification of two main hallmarks: extracellular neuritic plaques and neurofibrillary tangles formed by post-translational modified tau protein.
These limitations mean that it is possible to only make a probable AD diagnosis “based on clinical criteria, including medical history, physical examination, laboratory tests, neuroimaging and neuropsychological evaluation.”

The best known AD biomarker is beta-amyloid(1-42) in cerebrospinal (CSF) fluid – customarily designated as CSF Aβ(1–42). Its level in AD patients has been shown to be significantly reduced compared to controls, while levels of shorter Aβ(1–40) forms either remain unchanged or increase. Reduced levels of Aβ(1–42) are due to lower clearance of Aβ from the brain to CSF, as well as enhanced aggregation and plaque deposition. In addition, changes in CSF Aβ levels differ based on the disease.  Decreased levels of Aβ(1–38) correlate with frontotemporal lobe dementia (FTLD) while a reduction in Aβ(1–37) levels is associated with Lewy body dementia (LBD). 
CSF Aβ levels are, however, not considered to be reliable on their own. In June 2014, a meta-analysis on 14 studies and 1,349 patients by researchers at Imperial College London concluded that CSF Aß levels had “marginal clinical utility” and “cannot be recommended as an accurate test for Alzheimer’s disease.”
To ensure greater accuracy in diagnosis, several studies recommend combining CSF Aβ(1–42) concentrations with Aβ42/Aβ40 ratios. However, this has not been uniformly accepted, as some researchers found no evidence of the utility of Aβ42/Aβ40 ratios. 

Aβ42 and tTau
An alternative approach consists of combining CSF Aβ42 concentrations with another biomarker – total tau (tTau), which is a microtubule-associated protein. One study in Germany has suggested using both CSF Aβ42 and total tau as well as the Aβ42/Aβ40 ratio for diagnosing AD.
In healthy controls, levels of tTau increase with age, and some studies have sought to establish reference values for different age groups. tTau levels are measurably higher in AD patients as compared with age-matched control subjects and may be a prognostic marker for conversion from mild cognitive impairment (MCI) to AD. Studies have also found high CSF tau levels in 90% of MCI cases progressing at a later date to AD, but not in cases with stable MCI.

AD guidelines remain heterogeneous and in flux

The heterogeneous and evolving nature of diagnostic choices above is also echoed in the work of professional societies.
There are currently two sets of guidelines for the diagnosis of asymptomatic and symptomatic Alzheimer’s disease. One was published in 2007 by an International Working Group (IWG), while the second consists of recommendations in 2011 by the US National Institute on Aging and the Alzheimer’s Association (NIA–AA).
The NIA-AA criteria define three phases in the progression of AD: preclinical AD, mild cognitive impairment due to AD and dementia due to AD. NIA-AA highlights the need for significant additional research “to validate the application of biomarkers,” and acknowledges this is “likely to take more than a  decade to fully accomplish.”
Interpreting the two guidelines poses its own challenges. A comparison made by a team in the Netherlands “revealed differences in approach, terminology, and use of cognitive markers and biomarkers.” However, it found that patients “who meet the International Working Group criteria will also meet the NIA-AA criteria and vice versa,” and called for further research “to validate the criteria.

Biomarkers redefine AD
In 2010, one year before the NIA–AA recommendations, the IWG revised its definition of AD in order to provide “broader diagnostic coverage” of the disease’s clinical spectrum. The key reason for the revision was “access to reliable biomarkers in vivo” which had “radically changed” the definition of AD.”
In the coming years, it is clear that field trials will be “needed to establish whether the diagnostic criteria will work effectively in clinical or research situations.”

Clinical and research requirements
The difference between clinical and research situations is an important one. A key question for both clinician and researchers is whether the new criteria “are meant for research purposes or for clinical use.” The former are targeted at fields like drug testing with their validation paving the way for (new) clinical criteria. Clinical criteria “have to pass a higher bar in terms of robustness and accuracy.” 

Health authorities in the US and Europe have yet to formally qualify any AD biomarkers. Until then, most experts suggest that the IWG guidelines should be considered as research criteria, for use “in some drug trials and in academic medical settings that conduct clinical research.” The NIA/AA criteria, on their side, are published as a mixture of clinical- and research-grade, with the clinical portions of criteria meant for use in clinical settings, while “anything involving biomarkers and all of the preclinical AD criteria are for research use.”

Phospho-tau provides third dimension to tTau and Aβ(1–42)
As of now, it is accepted that the best method for diagnosing AD in patients is to measure CSF levels of Aβ(1–42) and tTau, along with a third biomarker phospho-tau (P-tau). Several studies have suggested “that abnormal hyperphosphorylation of tau in the brain plays a vital role in the molecular pathogenesis of AD.”
Tau is hyperphoshorylated at a potential total of 39 sites in AD. Its detection at position 181 and 231 in particular are “significantly enhanced in AD compared to controls.” Both have been shown to distinguish AD from controls and vascular dementia (VaD), frontotemporal lobe dementia (FTLD) and Lewy body dementia (LBD).
The combination of Aβ(1–42), tTau and P-Tau significantly increases diagnostic validity for sporadic AD, yielding “a combined sensitivity of >95% and a specificity of >85%. tTau and P-Tau have also been central to a major multi-centre trial in Europe, part of the European Alzheimer’s Disease Neuroimaging Initiative (E-ADNI).

Alternatives to CSF
AD diagnosis has been largely based on collecting CSF and this is accompanied by major limitations. CSF requires lumbar puncture, which is invasive and has many potential side effects. The routine screening of patients is therefore difficult. So too is their follow-up over several years.
Given this, researchers have sought to search for AD biomarkers in other body fluids. Saliva and urine are easily collected but “blood analysis is the gold standard.” Although the correlation of pathological changes in the brain to concentration of blood analytes remains unknown, a search for AD biomarkers in blood is likely to first target “accepted CSF markers, such as Aβ and tau-related biomarkers,” and then extend to factors involved in inflammation, protein ageing and cell death, and cerebrovascular dysfunctions. It seems very plausible that, at some point in the next few years, the combining of different blood-derived AD biomarkers leads to the definition of a patient-specific signature.
The pilot European trial on Alzheimer’s Disease Neuroimaging Initiatives (E-ADNI) measured both CSF and plasma-derived Aβ and found that higher diagnostic accuracy was obtained with frozen rather than fresh samples. E-ADNI also confirmed the feasibility of a multicentre AD biomarker programme for future clinical trials.

Microarrays and mass spectrometry
Meanwhile, researchers have been seeking to extend the search for other novel biomarkers.
One target is to use screening technologies such as microarrays and mass spectrometry, supported by bioinformatics. These would increase knowledge of disease-related changes in order to uncover novel AD biomarkers, open the way to quick and inexpensive diagnosis of AD and for gauging therapeutic relevance.
Some of the specific objectives here target the measurement of Aβ oligomers, to improve diagnostic specificity. A good example is surface-enhanced laser desorption/ionization-time-of-flight-mass spectrometry (SELDI-TOF-MS) which has emerged as an ideal method for the simultaneous detection and quantification of a variety of Aβ peptide cleavage products.,

Staying ahead
As disease modifying therapies based on new biomarkers are developed in clinical trials, it will be increasingly relevant to put the biomarkers to use. One way to do this is by accelerating the launch of interventions that arrest and (eventually) reverse AD.
The outlook is encouraging. A study recently published by researchers at Washington University in the US observes that clinicopathologic and more recent biomarker data suggest that AD pathology “begins to accrue approximately 10 to 20 years before any cognitive signs or symptoms”. This provides a window of opportunity for the initiation of secondary prevention trials that aim to prevent the development of symptoms in individuals while they are still cognitively normal.
Such steps were already foreseen by the NIA-AA criteria mentioned above. The guidelines note that if we can “definitively determine the risk of developing Alzheimer’s dementia in people who have biomarker evidence of brain changes but are not showing outward symptoms, we will open an important window of opportunity to intervene with disease-modifying therapies, once they are developed.