The Brain AT2R- A Potential Target for Therapy in Alzheimer’s Disease and Vascular Cognitive Impairment: A Comprehensive Review of Clinical and Experimental Therapeutics

Abstract

Dementia is a potentially avertable tragedy, currently considered among the top 10 greatest global health challenges of the 21st century. Dementia not only robs individuals of their dignity and independence, it also has a ripple effect that starts with the inflicted individual’s family and projects to the society as a whole. The constantly growing number of cases, along with the lack of effective treatments and socioeconomic impact, pose a serious threat to the sustainability of our health care system. Hence, there is a worldwide effort to identify new targets for the treatment of Alzheimer’s disease (AD), the leading cause of dementia. Due to its multifactorial etiology and the recent clinical failure of several novel Amyloid-β (Aβ) targeting therapies, a comprehensive “multi-target” approach may be most appropriate for managing this condition. Interestingly, Renin angiotensin system (RAS) modulators were shown to positively impact all the factors involved in the pathophysiology of dementia including vascular dysfunction, Aβ accumulation and associated cholinergic deficiency, in addition to Tau Hyperphosphorylation and insulin derangements. Furthermore, for many of these drugs, the pre-clinical evidence is also supported by epidemiological data and/or preliminary clinical trials. The purpose of this review is to provide a comprehensive update on the major causes of dementia including the risk factors, current diagnostic criteria, pathophysiology and contemporary treatment strategies. Moreover, we highlight the Angiotensin II receptor type 2 (AT2R) as an effective drug target and present ample evidence supporting its potential role and clinical application in cognitive impairment to encourage further investigation in the clinical setting.

INTRODUCTION

Nearly every minute someone in the US develops Alzheimer’s disease (AD) [1]. The incidence and prevalence of AD increase exponentially with age, essentially doubling in prevalence every 5 years after age 65 [1, 2]. In fact AD, which is almost always accompanied by vascular cognitive impairment (VCI), is the only leading cause of death that’s still on the rise [1, 3]. Between 2000 and 2018, deaths from heart disease have decreased while deaths from AD have significantly increased and are projected to increase even further in the coming years [1]. Moreover, total annual payments including direct and indirect health care costs, long-term care and hospice for people with AD and other dementias are projected to increase from $305 billion in 2020 to more than $1.1 trillion in 2050 [1]. In the US, such costs have already exceeded those of cancer and heart diseases [4]. The realization of its paramount public health impact has led the US government, to develop the “National Alzheimer’s Project Act” (Public Law 111–375) in an attempt to cope with AD and reduce its devastating effects [4].

Understanding Alzheimer’s disease and Vascular Cognitive Impairment

Alzheimer’s disease (AD)

Alzheimer’s disease is an age-associated, progressive, irreversible, multifactorial, neurodegenerative brain disorder. It begins many years before symptoms emerge and causes gradual cognitive decline, behavioral changes, psychiatric disturbances, dementia and eventual death from complete brain failure [1]. The majority (over 95 %) of AD patients, are diagnosed with “late-onset AD” (LOAD), also called “sporadic AD” (sAD). In these patients symptoms do not appear until after age 65, and the incidence continues to increase with age [5, 6].The remaining 5% of AD patients have “early-onset AD” (EOAD) or “Familial AD” (FAD). These patients carry rare genetic mutations to amyloid precursor protein (APP), presenilin-1 (PS-1), and presenilin-2(PS-2) that cause the initial symptoms to occur in as early as the person’s thirties [5]. Furthermore, some studies revealed that apolipoprotein epsilon four (Apoε4) allele also increases this risk [5].

The central histopathological hallmarks of Alzheimer’s disease are the deposition of misfolded amyloid-β (Aβ) protein aggregates in the form of senile plaques and the formation of neurofibrillary tangles (NFT), composed of hyperphosphorelated tau, with loss of synaptic connections in the brain [7]. These changes are accompanied by chronic neuroinflammation, oxidative stress as well as mitochondrial dysfunction, which ultimately lead to neurovascular degeneration and complete dementia [1, 7].

Although AD is the most common cause of dementia, accounting for an estimated 60–80 % of cases, it typically exists alongside a number of other causes of dementia that tend to have overlapping or related neuropathological processes. The most common of these is vascular cognitive impairment (VCI) [1]. In fact, the largest proportion of dementia cases have a “mixed” pathology, comprising features of AD (amyloid plaques and neurofibrillary tangles) as well as cerebrovascular characteristics, typical of VCI [4].

Vascular cognitive impairment (VCI)

Vascular cognitive impairment (VCI), which includes post-stroke cognitive impairment (PSCI), is the 2^nd leading cause of dementia in the world and covers between 25 and 30% of total dementia cases [8]. VCI defines alterations in cognition, attributable to cerebrovascular causes, regardless of pathogenesis or severity [4]. This can develop either as a result of a subclinical brain injury, as with long-term chronic hypoperfusion, transient ischemic attack or silent brain infarction (only seen with neuroimaging), or due to a clinically manifested injury as with an acute ischemic/ hemorrhagic stroke associated with single/ multiple infarcts. The severity of VCI can range from subtle deficits to overt dementia [9]. Cerebrovascular disease and AD frequently coexist [6, 7]. In fact, ischemic disease affects up to 90% of patients with AD, with major infarctions representing one third of vascular lesions and one third of putative cases of vascular dementia having coincidental pathological features of AD [10, 11]. Furthermore, studies confirm that plaques and tangles are more likely to cause AD if strokes or damage to the brain’s blood vessels are also presents [9, 12].

Frontotemporal dementia (FTD), Parkinson’s disease dementia (PDD), Lewy body dementia (LBD)

Other less common causes of dementia include frontotemporal dementia (FTD), Parkinson’s disease dementia (PDD) and Lewy body dementia (LBD). Frontotemporal dementia (FTD), which accounts for only 3% of dementia cases in those ≥65, may actually be among the most common cause of dementia in those younger than 60 [1]. This type of dementia is associated with a neural accumulation of transactive response DNA-binding protein (TDP-43) or tau. Unlike AD which is typically associated with memory loss (hallmark symptom), the main early features of FTD are language/speech difficulties and behavioral disturbances. Lewy body dementia (LBD), on the other hand, is associated with abnormal neuronal aggregations of an α-synuclein protein forming lewy bodies. The initial symptoms of LBD include sleep disturbances, clear visual hallucinations and visuospatial impairment. Although LBD accounts for ≈5% of those with dementia, most people with LBD also have AD pathology. Parkinson’s disease dementia (PDD) tends to develop after years of living with Parkinson’s disease. With this form of dementia, the α-synuclein protein aggregates occur in the substantia nigra, which is the “movement center” of the brain and result in degeneration of dopaminergic neurons. This causes motor symptoms (tremor, rigidity, loss of balance), which appear early on in the course of the disease, and may progress to cognitive impairment and dementia [1].

Major Acquired Risk factors

Hypertension and Diabetes are among the top modifiable risk factors for development and progression of cognitive impairment and dementia, including both VCI/PSCI and Alzheimer’s disease (AD) [8, 9, 13]. In fact, hypertension alone, which increases the risk of dementia both independently and by increasing the risk of stroke, can predict the development of dementia in nearly 60% of subjects with executive dysfunction [8]. Furthermore, longitudinal cohort studies have established a significant link between mid-life hypertension and AD [14].This is also true for diabetes which was shown to increase the risk of all type dementias by 73% [15]. Moreover, findings from the prospective cohort study, recently published in JAMA [16], showed that having two or more midlife vascular risk factors of hypertension, diabetes or dyslipidemia was significantly associated with elevated amyloid deposition (characteristic of AD) in the brain compared to none (61.2% vs 30.8%) [16]. This was further supported by results from the Harvard aging brain study which showed that vascular risks were associated with impending cognitive decline, both alone and synergistically with Aβ [12]. Another interesting finding is that smoking was associated with a 70% greater risk for developing AD, a risk that increases proportionally with cumulative cigarette exposure [17].

Diagnosis of AD and VCI

Alzheimer’s disease (AD)

Research currently identifies three stages of Alzheimer’s disease: preclinical AD, mild cognitive impairment (MCI) and Alzheimer’s dementia [1]. The first stage is only associated with pathological features while the last two stages, are also accompanied by symptoms of varying degrees [19]. With MCI there is evidence of lower performance in one or more cognitive domains, that is greater than one would expect for the patient’s age and educational background. Although the domain most often affected is episodic memory (i.e., the ability to learn and retain new information) it may also affect other cognitive domains, including executive function, attention, language and visuospatial skills. With Alzheimer’s dementia, the final stage of disease, the symptoms have progressed to the point where they impair a person’s ability to function independently. Note that the Alzheimer’s dementia phase is further subdivided into stages of mild, moderate and severe, according to the degree to which symptoms interfere everyday activities [1].

Alzheimer’s disease is fundamentally a clinical diagnosis. Cognitive impairment is detected and diagnosed through a combination of (1) history- obtained from the patient and a knowledgeable informant in addition to (2) an objective cognitive assessment and neuropsychiatric testing [19]. Numerous clinical instruments may be used to assess patients. These are rapid tests of memory and cognition that take up to 15 minutes to perform and include the mini-mental state exam (MMSE), the short blessed test (SBT) also called the Orientation-Memory-concentration Test and several others [20]. Additionally, routine blood tests may be used to rule out other causes of cognitive symptoms, such as tumors, endocrine issues or certain vitamin deficiencies [1].

Although the core clinical criteria for AD continue to be the cornerstone of diagnosis in practice, but imaging and cerebrospinal fluid (CSF) biomarker evidence may supplement the standard clinical presentation in certain instances [18]. These may be used to enhance the pathophysiological specificity and diagnostic accuracy, but only in specialized clinical research settings to monitor the effect of study treatments (facts and figures 2020) or under the Alzheimer’s association appropriate use criteria [18]. This is because further study is needed to refine, validate, and standardize imaging and biomarkers before they are ready for common clinical practice.

Vascular cognitive impairment (VCI)

While disturbances in episodic memory are considered a hallmark of pure AD, the salient feature of VCI is executive dysfunction, which includes impairments in attention, mental processing speed, working memory and cognitive flexibility.

To date, all diagnostic criteria to characterize cognitive syndromes associated with vascular disease are based on 2 factors:

1. Demonstration of a cognitive impairment affecting at least 1 cognitive domain, in the comprehensive cognitive/ neuropsychological battery for suspected VCI and

2. History of clinical stroke, or presence of vascular disease by neuroimaging [10].

Available treatments for AD and VCI

Alzheimer’s disease (AD)

There are only 5 medications currently approved by the U.S. Food and Drug Administration (FDA) for AD (Table I). Three of those rivastigmine [21], galantamine [22], donepezil [23] are cholinesterase inhibitors, while memantine [24] is an N-methyl-D-aspartate (NMDA) glutamate receptor antagonist. The newest formulation (Namzaric®), approved in 2014 for moderate-to-severe AD, combines memantine with donepezil for additive efficacy [25]. Unfortunately none of these medications cure, prevent, reverse, halt or even modify the progressive nature of the disease [1, 26, 27], They serve only as palliative measures, and their efficacy decreases over time [28]. Moreover, these medications are often associated with undesirable side effects including nausea, vomiting, hyperacidity, headache, insomnia, dizziness, and fatigue [21–23, 25].There is a dire need for alternate treatment modalities that would delay the course of this disease, prevent neurovascular degeneration and ultimately preserve cognitive functioning. A primary goal that forms the foundation of the US national plan for AD is to prevent and effectively treat AD by 2025 [6].

Vascular cognitive impairment (VCI)

Currently, no specific treatments for VCI/ PSCI have been approved by the US FDA. Nevertheless, Cochrane clinical trial reviews concluded that donepezil can be useful for cognitive enhancement in patients with vascular dementia (VaD), and galantamine for patients with mixed AD/VaD (Class IIa; Level of Evidence A). The benefits of rivastigmine and memantine are not well established in VaD (Class IIb; Level of Evidence A) [9].

Additionally, since hypertension is the single most important risk factor for both vascular and Alzheimer’s type dementias [29], treatment of hypertension received the highest evidence-based grade and is recommended for all those at risk for VCI (Class I; Level of Evidence A) [9]. Even in AD, which was once thought to be primarily caused by amyloid pathology, hypertension and associated vascular dysfunction, such as small vessel disease, were shown to be of greater importance than amyloid itself in terms of influencing the disease course, especially in older individuals. Therefore, modification of risk factors for small vessel disease is an important therapeutic goal for both AD and VCI [29, 30].

Pathophysiology

Although basic neuroscientific research has contributed, and continues to contribute, invaluable insight into disease mechanisms and pathophysiology [31], a major stumbling block has always been determination of the correct chronology of those factors and events that give rise to AD. Overall, there are a multitude of different players involved in the pathology of AD. These interact and integrate, through an intricate network of events, to maintain a perpetual state of homeostatic dysfunction, with chronic inflammation that gradually results in widespread cerebral damage and cognitive impairment. This is further complicated by the interaction of such factors, along with their resultant effects, on disease progression as well as neuropathological and clinical characteristics [11, 32]. Nevertheless, experts agree that, regardless of the precipitating event, or specific type, all forms of cognitive impairment/ dementia incorporate aspects of vascular dysfunction [33]. Note that, due to the multifactorial nature and neuropathological complexity of this disease; we will focus on its main components (Figure 1).

Figure 1.

A useful analogy describing what goes on in an AD brain is that of an inflammatory inferno. Aβ is fundamental in such pathogenesis, as it “fuels” the fire of neuroinflammation, analogous to dry tinder. It gradually accumulates and sets off a sequence of events, with neuroinflammation, oxidative stress and neurovascular damage all leading to cognitive impairment and dementia, similar to an uncontrollable forest fire with neurons for trees. 1) Aβ monomers aggregate to form oligomers, protofibrils, fibrils and eventually plaques. 2) Aβ oligomers activate microglia by binding to their surface RAGEs. These activated microglia release vast amounts of damaging proinflammatory mediators, (including IL-1β, TNF-α, NFκB) and free radicals (like O₂^.), which promote oxidative stress and further amplify inflammation. The escalating oxidative, pro-inflammatory environment promotes NVU injury. It results in 3) progressive endothelial atrophy along with pericyte and astrocyte damage, resulting in BBB disruption. 4) This toxic milieu also damages oligodendrocytes, resulting in demyelination and progressive axon loss. 5) Aβ accumulation also results in tau hyperphosphorylation, which leads to destabilization of microtubules, preventing axonal transport/trafficking of neurotransmitters to synapses, inhibiting impulse transmission. Hyperphosphorylated tau also tends to aggregate and form NFTs, which accumulate in neuronal cell bodies, leading to neurotoxicity and cell death.

Excessive Ang II mediated activation of AT1R causes vasoconstriction, which results in cerebral hypoperfusion and hypoxia, with reduced clearance of Aβ and increased inflammation. Ang II mediated activation of AT2R on the other hand results in vasodilation, improved cerebral blood flow, enhanced Aβ clearance with reduced inflammation and oxidative stress. ARBs were shown to preserve cognition both 1) directly by blocking AT1Rs and preventing their damaging effects and 2) indirectly by allowing an unopposed activation of the neuroprotective AT2Rs by the unbound Ang II hence promoting its beneficial effects. AT2R agonists like C21 act by directly activating the AT2R, hence facilitating Aβ clearance and reducing inflammation.

AD: Alzheimer’s disease, Aβ: Amyloid-β, Ang II: Angiotensin II, ARBs: AT1R blockers, AT1R: Angiotensin II receptor type 1, AT2R: Angiotensin II receptor type 1, BBB: Blood brain barrier, IL-1β: interleukin-1β NFκB: Nuclear factor kappa B, NFTs: neurofibrillary tangles, NVU: Neurovascular unit, OD: oligodendrocyte, RAGEs: receptors for advanced glycation end products, TNF-α: Tumor necrosis factor alpha

Vascular dysfunction-The Vascular hypothesis

Vascular dysfunction, which is now considered to be the initial trigger for cognitive impairment and AD, is tightly linked to neuronal dysfunction [32, 34]. In fact, the state of vascular insufficiency, hypoperfusion and endothelial dysfunction, that characterizes chronic conditions like hypertension and diabetes, induces an oxidative, proinflammatory environment that wreaks havoc on the brain [4]. Moreover, cerebral hypoperfusion is one of the earliest clinical manifestations of both the sporadic and familial forms of AD [35]. This is further intensified with age and typically results in impairments in brain metabolism (glucose and oxygen utilization), progressive blood-brain barrier (BBB) breakdown with demyelination, synaptic defects and disruption of the trophic coupling between neurovascular unit (NVU) components [4, 36].

Amyloid-β synthesis and processing- The amyloid cascade hypothesis

Amyloid-β (Aβ) is a ~4 kD peptide produced by the “amyloidogenic pathway” [37]. This generally involves sequential cleavage of amyloid precursor protein (APP), a type I transmembrane glycoprotein, by the β- secretase 1 (BACE1) enzyme followed by the γ- secretase enzyme complex. The initial cleavage by BACE1 yields an intermediary, membrane-bound, carboxy-terminal fragment of 99 amino acids (C99). This C99 can either be transported into the cell’s nucleus, to induce expression of apoptotic genes and promote cell death [38–40] or it can be further cleaved by presenilin, the catalytic subunit of γ- secretase to release the toxic Aβ peptide, a 40–42 amino-acid fragment, into the extracellular environment [41].

Aβ may exist as a heterogeneous mixture of peptides with different solubility, stability as well as biological and toxic properties [38]. It can aggregate to form oligomers, protofibrils, fibrils, or eventually plaques, which are one of the pathological hallmarks of AD [38, 41]. Contrary to what was previously thought, it is actually the small soluble Aβ oligomers, also referred to as Aβ derived diffusible ligands (ADDLs) that are reportedly more neurotoxic than the insoluble fibrils (plaques). These ADDLs assemble under conditions that block fibril formation and can destroy hippocampal neurons at nanomolar concentrations [38]. These toxic oligomers also bind to α−7 nicotinic acetylcholine (ACh) receptors, diminishing the release of Ach [11, 26] and impairing synaptic transmission [38] in addition to inhibiting critical neuronal activities, including long-term potentiation (LTP), synaptic plasticity and memory processing [38].

Under normal physiologic conditions Aβ synthesis is minimal with any excess rapidly eliminated by transvascular transport. Instead, APP is processed by the “non-amyloidogenic pathway” which involves cleavage by α- secretase, to yield a soluble extracellular fragment (sAPPα). This regulates neural excitability and enhances synaptic plasticity, learning and memory, in addition to boosting neural resistance to oxidative and metabolic stress [39, 40, 42]. The amyloid cascade hypothesis proposes that Aβ accumulation in the brain, which results from an imbalance between its generation and clearance, is fundamental in AD pathologenesis [43]. This is because Aβ accumulation sets off a sequence of events, most notably neuroinflammation, vascular damage and tau Hyperphosphorylation . With a loss of synapses in addition to neuronal dysfunction and death, that all lead to cognitive impairment and dementia [39].

Amyloid-β effects on the neurovascular unit NVU

In addition to its well established neurotoxic effects, Aβ negatively affects every other component of the NVU. It activates surrounding microglia by binding to their surface receptors for advanced glycation end products (RAGEs), stimulating the release of damaging chemokines and pro-inflammatory cytokines, including interleukin-1β (IL-1β), IL-6, Tumor Necrosis Factor-α (TNF-α) and interferon-gamma (IF-γ). These inflammatory mediators not only promote neurovascular injury and cholinergic dysfunction directly, they also recruit additional inflammatory mediators, like NFκB-which further aggravate inflammation by sustaining microglial activation [11, 43]. This chronically activated microglia amplify oxidative stress by generating free radicals and glutamate, the latter which induces excitotoxicity and increases cytosolic Ca²⁺ to trigger apoptosis. In fact, Aβ itself triggers apoptosis by disrupting mitochondrial function, impairing oxygen consumption and preventing ATP production. This increases superoxide radical formation, causing oxidative stress and release of cytochrome c, leading to cell death [11, 28, 44]. Furthermore, the escalating oxidative, pro-inflammatory environment results in progressive endothelial atrophy along with pericyte damage and microvascular rarefaction. The ensuing BBB disruption results in increased permeability and edema that compresses cerebral microvessels, further reducing cerebral blood flow and worsening tissue hypoxia. This toxic milieu damages oligodendrocytes, which also rely on ECs for trophic support. Oligodendrocyte damage results in axonal demyelination and loss of energy-saving saltatory conduction, increasing metabolic demands and local energy deficit. In addition to the brain dysfunction caused by slowing of action potential transmission, demyelination threatens axonal integrity and increases their vulnerability by exposing them to the deleterious effects of cytokines and free radicals, present in such hypoxic environment eventually leading to complete axonal loss [45]. In addition to amplifying oxidative stress and neuroinflammation, Aβ is also a potent vasoconstrictor [40] that reduces its own transvascular clearance, leading to additional accumulation and toxicity that create a vicious cycle.

Amyloid-β effects on Tau

Tau is a soluble microtubule associated phosphoprotein (MAP), normally confined to axons [42, 46]. It promotes the polymerization and assembly of tubulin into microtubules, which are the structures necessary for axonal transport and trafficking of neurotransmitters to synapses. It also plays an important role in microtubule stabilization, dynamics and overall function within neurons [47]. The level of Tau phosphorylation regulates microtubule binding, with hyperphosphorylated tau lacking affinity for microtubules. Therefore, hyperphosphorylation of tau is highly detrimental as it leads to its detachment from and destabilization of microtubules [48]. Hyperphosphorylated tau also tends to self-associate, sequester normal tau and aggregate into insoluble paired helical filaments (PHFs) that form neurofibrillary tangles (NFTs). These NFTs accumulate in neuronal cell bodies, as well as synapses, and cause neurotransmitter deficits and eventual neuronal death [28]. Tau hyperphosphorylation is mainly a result of Aβ accumulation [49] In fact, a causal relationship has been established, linking Aβ accumulation with tau hyperphosphorylation, aggregation and associated cytoskeletal derangements [11].

Insulin-Signaling Pathway

Another metabolic disturbance of emerging importance in Alzheimer’s disease and VCI involves insulin signaling in the brain, which ties into synaptic function and energy homeostasis [44]. Deficits in brain insulin signaling in AD have been recognized for years [50], even adding the notion that AD should be regarded as a brain form of diabetes, in which insulin resistance and deficiency develop either primarily in the brain, or due to systemic insulin resistance with secondary brain involvement [5, 51]. In fact, several experimental models have shown that brain insulin resistance, or deficiency, impair learning and memory [52–54]. Normally, insulin and insulin-like growth factor-1 (IGF-1) initiate signals needed for neuronal survival, plasticity, and metabolism in the brain [51]. This takes place by their activation of the phosphatidylinositol-3-kinase (PI3K)–Akt and the mitogen-activated protein kinase–extracellular signal-regulated kinase (MAPK/ERK) pathways. Insulin resistance and glucose intolerance, common among those with advanced AD [55], impair PI3K-Akt signaling [53]. This not only renders neurons energy-deficient and impairs synaptic plasticity but it is also associated with an up-regulation and activation of the tau kinase glycogen synthase kinase 3β (GSK-3β) with increased levels of phospho-Tau and Aβ42 [51]. In fact, insulin resistance and impaired PI3K-Akt signaling with increased activation of GSK-3β are common features of AD in human brains [51, 55]. Insulin resistance also results in elevated serum glucose levels, which directly damage hippocampal structures [56] and lead to a rise in advanced glycation end products (AGEs) [57]. These AGEs activate their respective receptors, RAGEs, to increase oxidative stress, hence exacerbating vascular inflammation, thrombosis and vascular damage [57].

Brain renin angiotensin system (RAS) and its link to cognition

Angiotensin II subtype 1 receptors (AT1R) Hyperactivation

The intrinsic brain renin angiotensin system (RAS) plays an essential role in learning and memory [58, 59]. Angiotensin II (Ang II), a hormone and neuropeptide produced mostly in CNS glial cells by angiotensin converting enzyme (ACE) mediated hydrolysis of angiotensin I, is the central RAS effector [27]. This neuropeptide appears to be actively involved in the pathogenesis of AD and is responsible for triggering neurovascular damage and dysfunction through “hyperactivation” of its angiotensin II subtype 1 receptors (AT1R) pathway [27, 60]. The increased AT1R activity results in neurovascular inflammation, with impaired cerebrovascular autoregulation and accentuated vasoconstriction. This leads to cerebral hypoperfusion and hypoxia, the latter which facilitates production of Aβ by increasing APP transcription and activating β and γ -secretase enzymes, essential for Aβ synthesis [27]. As was mentioned before, Aβ is associated with a number of neurotoxic effects. Overall, despite the diversity of underlying causes, the neurovascular alterations have similar mechanistic basis, with vascular insufficiency and hypoperfusion paving the way for neuronal dysfunction, injury and cognitive impairment.

Angiotensin II subtype 2 receptor (AT2R) Modulation

The other major RAS pathway involves stimulation of angiotensin II subtype 2 receptors (AT2R). Accumulating evidence indicates that this arm of the RAS is neuroprotective, as it promotes vasodilation, improves cerebral microcirculation, reduces oxidative stress, decreases inflammation (both by reducing expression of proinflammatory cytokines and increasing expression of anti-inflammatory mediators) and protects against brain damage [59]. In fact, stimulation of the AT2R has emerged as a novel therapeutic strategy in vascular and central nervous system diseases by virtue of its anti-inflammatory, antiproliferative and tissue regenerating properties [61–63]. Compound 21 and CGP42112A are the direct AT2R agonists currently available for research [61].

AT2-receptor agonist CGP42112A

CGP42112A, developed by Ciba-Geigy [64], is merely an experimental tool as it provides proof of principal for AT2R mediated neuroprotection but has several important limitations. Firstly, CGP42112A is a partial AT2R agonist with high binding affinity but low potency, so while occupying the receptor at relatively low concentrations, it requires much higher concentrations to actually stimulate it, making the results difficult to interpret, since the turning point between antagonism and agonism is not clearly defined [64]. Also, in addition to requiring higher concentrations to stimulate AT2R, it will tend to lose its AT2R selectivity at higher doses, making it even more challenging to work with [64, 65]. Moreover, CGP42112A is given to animals by intracerebroventricular (ICV) injection, which is not practical in the clinical setting. and it is a peptide that is rapidly degraded in vivo, rendering it unsuitable for use in humans [66].

Compound 21(C21/M024)

Compound 21 (C21), is a selective, orally active, highly soluble direct nonpeptide AT2R agonist developed by Vicore Pharma (Göteborg, Sweden) [67]. It is hydrophilic and rapidly absorbed with median Tmax at 40 min in humans and 90 min in rats, an oral bioavailability of 20 –30% and a half-life (t_1/2) of 4 and 5.4 hours (h) in rats [68] and humans [69] respectively. Owing to its favorable physicochemical and pharmacokinetic properties, C21 is much more clinically relevant than CGP42112A [67]. Moreover, doses of up to 100 mg were found to be safe and well tolerated in human subjects [63]. Treatment with C21 has been shown by multiple groups to have positive effects on cognition and memory [34, 36, 60, 62, 65, 117, 127, 129]. It not only reduced cognitive decline in various animal models of VCI, but it also enhanced spatial memory in both normal and ICV Aβ injected mice [70]. In fact, our group has shown that C21, when administered chronically, effectively preserved overall cognitive function, including non-spatial recognition and short-term working memory, associative learning, spatial reference and long – term memory as well as cognitive flexibility [34, 36, 60]. This cognitive preservation was apparent in young [60] and aged hypertensive [34] as well as normotensive animals, post-ischemic insult [36], with no effect on blood pressure [60, 69].

The proposed mechanisms of AT2R mediated cerebrovascular protection and cognitive preservation by C21 include an increase in cerebral blood flow and reduction in Aβ accumulation, with the suppression of chronic reactive microgliosis and neuroinflammation [36, 60, 62].This is essential, considering that neuroinflammation is an early and consistent feature of AD [71]. In fact, the three signaling pathways known to mediate neuroinflammation, namely the JAK/STAT pathway [72] NF-kB signaling pathway [73] and COX2 synthesis pathway [74] are all inhibited by AT2R stimulation [67]. This AT2R stimulation by C21 also increased neuronal levels of VEGF [75] and BDNF [76], both neurotropic factors essential for overall health and survival of neurons. This drug also displayed vascular protection and was shown to increase occludin, claudin-5 and ZO-1 expression, hence preserving BBB integrity [77]. It was also shown to improve insulin sensitivity (attenuate insulin resistance) and decrease oxidative stress, partly by enhancing the expression and overall activity of PPARγ [70].

Angiotensin II subtype 1 receptor (AT1R) receptor blockers (ARBs)

There are eight ARBs currently used in the clinical setting (Table II). Although most ARBs share the same characteristic pharmacophore, imidazole derivative, with a common class effect the group is chemically heterogeneous [78]. Differences in their physiochemical properties, pharmacokinetic profiles and physiological effects are due to different substituents [78]. Nevertheless, each member in the class has at least one study documenting evidence of positive impact on cognitive function, which points to a class effect when it comes to cognitive benefits. These agents are both vascular and neuroprotective [79]. In fact, they were shown to reduce the cognitive impairments associated with vascular disease, AD and other neurodegenerative conditions [80, 81].

We and others have demonstrated that the positive effect of ARBs on cognition is two-fold. These agents work both 1) directly, by selectively blocking AT1R, hence inhibiting the detrimental pathway and 2) indirectly, by allowing an unopposed stimulation of the neuroprotective AT2R pathway by the unbound Ang II, hence promoting beneficial effects [60, 82]. This results in improved CBF, reduced Aβ toxicity, reduced oxidative stress, diminished vascular and tissue inflammation, in addition to enhanced cerebrovascular autoregulation/ function, increased resistance to ischemia, and inhibition of neurodegeneration [67]. Moreover, ARBs were also shown to effectively restore insulin mediated PI3K/Akt signaling [53], known to be defective in AD patients [83]. Several clinical and experimental studies showed that these and other RAS modulators improve insulin sensitivity and even reduce the risk and or incidence of type 2 diabetes, a major AD risk factor [84–86].

Medication Repurposing and potential therapies

In general, a new drug requires an average of 15 years to reach the market today and approaches nearly 2.6 billion dollars in research and development, before this new molecule can be approved and marketed [87]. The majority of new molecules never even make it to the market despite all the time and cost invested in its development. A way to minimize much of this lost effort and resources as well as substantially reduce cost is through medication repurposing.

Medication repurposing, also referred to as medication repositioning is defined as “the application of established drug compounds to new therapeutic indications” [14]. A repurposed drug is already licensed, has been FDA approved, for an initial indication, so has already passed many of the initial steps including in vitro and pre-clinical screening, chemical optimization, toxicological studies and bulk manufacturing in addition to formulation development. It has also shown potential clinical efficacy for another “proposed” indication, as evidenced by epidemiological cohort studies, open-treatment studies and preliminary clinical trials, enabling it to advance to the market, for the proposed indication, much quicker [14].

Indeed, drug repositioning has resulted in therapeutic success in many areas including cancer and cardiovascular disease [14]. Moreover, a substantial body of clinical and experimental evidence indicates that drugs targeting the renin angiotensin system, mainly ARBs, are associated with preservation of cognition and work to improve cerebral function and Alzheimer’s-like brain pathology to varying degrees. These studies will be detailed in the next section and summarized in Tables III and IV, for animal and clinical studies respectively.

Experimental evidence-animal studies

There is abundant pre-clinical evidence supporting the efficacy of ARBs, like candesartan, in several types of dementia. In fact candesartan[88] alone showed ample benefits in various models of cognitive impairment [60, 89–95]. It effectively preserved cognitive function in young [60] and aged [36] hypertensive animals post-ischemic insult, even when initial administration was delayed [36, 60]. Some even founds that, the degree of improvement was more pronounced in older vs younger animals [91]. These cognitive benefits of candesartan went far beyond its effect on BP, as these were also seen with low sub-hypotensive doses [60, 89, 90, 95–97]. This was also true with azilsartan [65, 98] telmisartan [99–102] and olmesartan [103–105] all of which effectively ameliorated the cognitive decline, limited BBB disruption and significantly reduced oxidative stress and inflammation in animal models of VCI, without affecting BP. Candesartan also prevented cognitive decline associated with laporascopic surgery [96] as well as drug-induced cognitive impairment, resulting from streptozotocin [94], AngII [89] or anticholinergic agents like scopolamine [97], as was the case with valsartan [108–112] and losartan [108, 113, 114], both of which also prevented cognitive impairments resulting from other agents namely aluminum trichloride/ D–galactose [115] and sodium valproate [116]. Moreover, candesartan prevented the cognitive deterioration resulting from a traumatic brain injury [93, 117] as well as that induced by chronic stress [90] and diabetes [95]. Such benefits were also seen with telmisartan [118, 119] and olmesartan, which also effectively inhibited diet induced cognitive impairment [120]. Additionally, ARBs including candesartan, losartan, olmesartan and telmisartan successfully improved cognitive outcomes in various transgenic models of AD [92, 105, 121–124]. These agents reduced amyloid burden [105, 111, 124, 125] oxidative stress [123], inflammation and neuronal loss [92, 126], while also restoring cerebrovascular reactivity and BBB function [123] in transgenic AD mice [92].

Interestingly, the positive effects of ARBs like candesartan on memory, CBF, ACh level, and oxidative stress were blunted by concomitant blockade of AT2R, indicating that the AT2R contributes to these favorable effects [97]. The beneficial effects of unopposed AT2R stimulation were also confirmed with direct AT2R agonists CGP42112A and C21.

The AT2R agonists, C21 and CGP42112A showed positive effects in animal models of cognitive impairment [36, 60, 65, 117, 127]. These benefits, which were apparent in animals of both sexes (unpublished data), were not apparent in AT2R KO mice and were attenuated in wild type mice given the AT2R blocker, PD123319 [62], which confirms the requirement of AT2R for their positive action. Although CGP42112A is merely an experimental agent, intended for proof of concept studies, it improved functional recovery and cognitive performance in animals subjected to traumatic brain injury (TBI) [117]. This was accompanied by a dose-dependent reduction in lesion volume and enhanced neurogenesis [117]. The more recently developed AT2R agonist, C21, was associated with multiple benefits. This drug effectively preserved cognitive function and prevented the development VCI [34] and PSCI [36, 60] in young hypertensive as well as aged animals post-stroke [34, 36, 60]. It also markedly reduced cytotoxicity and cortical accumulation of Aβ₄₂ [60], preserved CBF and reduced inflammatory cytokine expression in BCAS induced cognitive impairment [128]. C21 also showed synergistic effects in preventing diabetes-induced cognitive impairment when given with the AD drug memantine [129].

Clinical evidence-human studies

A multitude of clinical trials, illustrated in Table IV support the benefits of RAS modulators in improving cognitive function and preventing dementia in various subsets of patients. This was evident in double-blind randomized controlled studies [110, 130–134], large, multinational open-label observational and longitudinal studies [135–138], prospective cohort analyses, [29, 139–141], meta-analyses [142,143] and retrospective studies [144–147].

In fact, RAS modulators (mainly ARBs) were shown to prevent cognitive and functional declines in older adult and were associated with a significantly lower incidence and progression of AD and dementia, compared to other agents [130, 132, 139, 144, 148, 149]. Indeed, RAS modulators were shown to be superior to other antihypertensive medications in preventing early declines in executive and overall cognitive function [143, 150]. This was true even among elderly diabetic patients without hypertension [139], those with hypertension but no prior cerebrovascular disease [142] or dementia [138, 151, 152] as well as in those already diagnosed with dementia [29, 139] or APOε4 carriers, at risk of cognitive decline and dementia [153]. Even in the case of better BP control, seen in non-RAS medication users, RAS users demonstrated slower declines in memory and attention and were a lot less likely to convert to AD than non-RAS users [150]. This indicates a specific benefit that went far beyond their actions on blood pressure [150] and was confirmed across the different sexes, races and ethnicities [154]. Although some found that the slowing of cognitive decline was more pronounced with BBB-crossing RAS medications [147,150,155], others found this to be independent of RASs’ lipophilicity or ability to cross the BBB [153].

Summary, future directions and clinical implications

Dementia has baffled the scientific community for centuries and research continues to explore innovative treatments and therapeutic strategies in an attempt to slow the course of its progression. Exactly what precipitates its insidious development remains a mystery, with its elusive nature and neuropathological complexity; one might expect that there is no “simple fix”. Nevertheless, initiating therapy in its early stage, using a class, of agents that target the multiple molecular pathways involved in its pathology, may prove effective in suppressing its otherwise inevitable progression.

Although RAS modulators alone may not “cure” dementia, there is substantial and reasonably consistent evidence supporting their benefit and potential application in management of AD and VCI, with additional clinical trials underway (Table V). These agents, which owe much of their cognitive benefit to the indirect activation of AT2R, may be good candidates for prevention of neurodegeneration and dementia in patients at high risk. This is because they effectively target the multitude of different factors involved in the pathophysiology of dementia. RAS modulators, namely ACEIs and ARBs, are also first line for individuals with comorbid conditions including hypertension and diabetes, kidney disease, heart failure or history of stroke [156]. These comorbid conditions are particularly common among older individuals with, or at high risk of, AD/VCI [12]. All these criteria make RAS modulators especially attractive, also given their extensive use, ease of administration (oral dosage, good pharmacodynamic characteristics), their great tolerability and favorable side-effect profile, in addition to their reasonable cost. Indeed, all of these agents, with the exception of C21 which is currently in the trial phase, are already FDA approved and marketed, so can be repurposed for AD/VCI as part of a comprehensive therapeutic plan/regimen. This repositioning would be an effective albeit lucrative option, given the lack of success with Aβ targeting therapies, which cost billions of dollars in drug development, formulation and testing yet failed to show benefit in clinical trials [14, 26, 28]. Nevertheless, future trials should also focus on selection of optimal doses of individual drugs within the class, as these may differ somewhat from those commonly used in the treatment of the other conditions, for which the class is indicated.

Another auspicious approach may be to employ the direct AT2R agonist, C21. This drug appears to be a good candidate for further investigation in clinical trials, since its use for prevention of dementia is well supported by preclinical and experimental evidence [34, 36, 60, 62].

This drug may be a valid alternative for preserving cognition in various subsets of patients, who may not be good candidates for therapy with other RAS modulators due to allergies or intolerance. Other suitable candidates for C21 may also include elderly normotensive patients with mild cognitive impairment, or those with heart valve disorders, severe carotid occlusive disease or atrial fibrillation and orthostatic hypotension, for whom extensive BP lowering may be detrimental. This would exploit the numerous protective benefits associated with AT2R stimulation, and hence sustain cognitive function, without affecting BP [60]. Although the selective AT2R agonist C21 may seem like the ideal candidate, with its multitude of potential clinical applications in addition to its encouraging data and supportive evidence in preclical models of AD and VCI, it is still far from ready to enter the market. This is because there are still several missing links, and with clinical trials currently at phase 1 it is unclear when and whether it will be available.

Future trials should also focus on identifying patients in the very early pre-symptomatic stages, using effective yet relatively simple diagnostic techniques. This is because there is often a delay in the development of symptomatic cognitive decline. This “preclinical phase” is likely the optimal stage for application of interventions to preserve cognition, as it would target their therapeutic time window of efficacy.

Unfortunately, most randomized clinical trials tend to be carried out on symptomatic patients, at which point the disease process has likely already progressed into the advanced, perhaps irreparable stages for interventions to be successful [14].

Indeed, with such a complex condition as dementia, a systematic approach must be employed for screening of patients in the early stages, as with the ongoing rrAD [157] and CEDAR [158] trials and instigating therapy, within the appropriate time window of efficacy. Ideally, a specialized set of regimens must be established, as part of a complete set of clinical guidelines and recommendations, for therapeutic management of patients in the various stages of disease. These should be continuously revised and updated as necessary to account for the most recent findings.

Abbreviations:

ACE

Angiotensin Converting Enzyme

ACEIs

Angiotensin Converting Enzyme Inhibitors

Ach

Acetylcholine

AChE

Acetylcholine esterase

AD

Alzheimer’s disease

ADAS-cog

Alzheimer’s Disease Assessment Scale-Cognitive

AH

Antihypertensive

Ang

angiotensin

APOε4

Apolipoprotein epsilon four

APP

Amyloid precursor protein

ARBs

AT1R blockers

AT1R

Angiotensin II receptor type 1

AT2R

Angiotensin II receptor type 1

Aβ

Amyloid-β

BACE1

β-secretase 1

βb

β-blockers

BBB

Blood Brain Barrier

BCAS

Bilateral Common Carotid Artery Stenosis

BCCAO

Bilateral common carotid artery occlusion

BDNF

Brain derived neurotrophic factor

BP

Blood Pressure

C21

Compound 21

CBF

Cerebral blood flow

CCBs

Calcium Channel Blockers

CCH

Chronic cerebral hypoperfusion

CCI

Controlled cortical impact

CDR

Clincal dementia rating

CGP42112A

peptide AT2-receptor agonist

CSF

cerebrospinal fluid

CV

Cardiovascular

DBP

Diastolic blood pressure

d-MCAO

distal middle cerebral artery occlusion

EPC

Endothelial progenitor cell

FAD

Familial Alzheimer’s disease

FDA

Food and drug administration

GEMS

Ginko evaluation of memory study

HCTZ

hydrochlorothiazide

HSCD

high-salt, high cholesterol diet

ICV

intracerebroventricular

IGF-1

insulin-like growth factor-1

IL-1β

interleukin-1β

IN

Intranasal

IP

Intraperitoneal

M1:M2 ratio

pro-inflammatory to aniinflammatory; macrophages

MCI

mild cognitive impairment

MCP-1

Monocyte chemoattractant protein-1

MMP

matrix metalloproteinase

MMP-9

matrix metalloproteinase-9

MMSE

Mini-Mental State Examination

MoCA

Montreal Cognitive Assessment

MRI

magnetic resonance imaging

NFT

Neurofibrilar tangle

NFκB

Nuclear factor kappa B

NMDA

N-Methyl-d-aspartate

NVU

Neurovascular unit

P- Tau

phosphorelated tau

PI3K

Phosphatidylinositol-3-kinase

PO

Per oral

PPARγ

peroxisome proliferator-activated receptor gamma

PROBE

Prospective Randomized Open, Blinded End-point

PSCI

post stroke cognitive impairment

RAS

Renin Angiotensin System

RCT

Randomized controlled trial

SBP

Systolic blood pressure

SHRs

spontaneously hypertensive rats

SHRSP

stroke prone spontaneously hypertensive rats

SQ

subcutaneous

STZ

Streptozotocin

T2D

Type 2 diabetes

TBI

Traumatic Brain Injury

t-MCAO

transient Middle Cerebral Artery Occlusion

TNF-α

Tumor necrosis factor alpha

TrkB

Tropomyosin receptor kinase B

T-Tau

total tau

UCCAO

Unilateral common carotid artery occlusion

VaD

vascular dementia

VCI

vascular cognitive impairment

VEGF

vascular endothelial growth factor

WMH

white matter hyperintensity

WT

wild type

Table Appendices

Appendix A:

Table I-.

FDA Approved Drugs for Alzheimer’s disease

Drug (Brand name) Manufacturer	Class	Initial FDA Approval	Indication	Off label use	Reference
Donepezil (Aricept®) Eisai/Pfizer.	Acetylcholine sterase (AchE) inhibitor	1996	Mild, moderate & severe AD	VaD (Class IIa; Level of Evidence A)	(ARICEPT) [23]
Rivastigmine (Exelon®) Novartis Pharmaceuticals	AchE inhibitor	2000	Mild-moderate AD		(EXELON) [21]
Galantamine (Razadyne®) APOTEX INC	AchE inhibitor	2001	Mild-moderate AD	Mixed AD/VaD (Class IIa; Level of Evidence A)	(RAZADYNE) [22]
Memantine (Namenda®) AJANTA PHARMA	N-methyl-D-aspartate (NMDA) receptor antagonist	2003	Moderate-severe AD		(NAMENDA) [23]
Memantine + donepezil (Namzaric) Allergan	NMDA receptor antagonist and AchE Inhibitor	2014	Moderate-severe AD		(NAMZARIC) [25]

Appendix B:

Table II-.

Angiotensin II Receptor 1 Blockers

Drug (Brand name) manufacturer	Initial FDA Approval	Indications	Pharmacokinetic profile (biological half-life/ oral bioavailability)	References
Losartan (Cozaar) Merck Sharp & Dohme Corp.	1995	Hypertension- Treatment Stroke- risk reduction* Diabetic nephropathy- Treatmentǂ	6–9 h/33%	COZAAR [114]
Valsartan (Diovan) Novartis Pharmaceuticals Corporation	1996	Hypertension- Treatment Heart Failure- Treatment Post-Myocardial Infarction- CV risk reduction	6 h/25%	DIOVAN [112]
Candesartan (Atacand®) AstraZeneca	1998	Hypertension- Treatment (adults and children) Heart Failure- Treatment	9 h/15%	ATACAND [88]
Telmisartan (Micardis) Boehringer Ingelheim Pharmaceuticals, Inc	1998	Hypertension- Treatment CV risk reduction in those unable to take ACEIs	20 h/42–58%	MICARDIS [100]
Eprosartan (Teveten) AbbVie Inc.	2001	Hypertension- Treatment	5 h/13%	TEVETEN [106]
Irbesartan (Avapro) Sanofi Company	2002	Hypertension- Treatment Diabetic nephropathy- Treatment ǂ	11–15 h/70%	AVAPRO [107]
Olmesartan (Benicar) Daiichi Sankyo, Inc.	2002	Hypertension- Treatment	14–16/29%	BENICAR [103]
Azilsartan (Edarbi) Takeda Pharmaceuticals America, Inc.	2011	Hypertension- Treatment	11 h/60%	EDARBI [98]

^*For Patients with Hypertension and Left Ventricular Hypertrophy

^ǂFor Patients with T2DM and Hypertension

Appendix C:

Table III-.

Animal studies documenting the effects of ARBs/AT2R agonists on cognition

1.Inhibition of cognitive decline in mice fed a high-salt and cholesterol diet by the angiotensin receptor blocker, olmesartan [120].
Animals	8 wk old male C57BL6 fed a high-salt, high cholesterol diet (HSCD) vs normal diet (ND)
Model	Diet induced cognitive impairment
Treatment/Intervention	PO olmesartan (3 mg/kg/d) or vehicle administered from age 8 wks – age 12 wks
Results/Conclusion	HSCD in mice was associated with significant learning impairment and cognitive decline. Olmesartan inhibited HSCD induced cognitive decline and significantly improved cognitive function to a level similar to that of ND mice. It also reduced BP and superoxide anion production as well as improved serum cholesterol and glucose concentrations.

2.Amelioration of cognitive impairment in the type-2 diabetic mouse by the angiotensin II type-1 receptor blocker candesartan [95].
Animals	8 and 15 wk old male diabetic (KK-Ay) and WT (C57BL6) mice
Model	Diabetes induced cognitive impairment
Treatment/Intervention	Candesartan at 2 different “nonhypotensive” doses (0.001% or 0.005%) given in chow at libitum Short-term treatment Initiated at age 15 wks (continued for 5 wks) Long-term treatment Initiated at age 8 wks (continued for 7 wks)
Results/Conclusion	Long-term Candesartan treatment markedly improved cognitive function to a level similar to that of age-matched controls. The lower dose of Candesartan (0.001%) starting from 8 wks of age also significantly restored cognitive function at 7 wks post treatment. Short-term administration of Candesartan (0.005%) starting from 15 wks of age inhibited the inevitable cognitive decline seen in untreated KK-Ay mice, with no apparent change in BP

3.Valsartan lowers brain β-amyloid protein levels and improves spatial learning in a mouse model of Alzheimer disease [111].
Animals	6 mo old WT and AD (Tg2576) mice
Model	Transgenic AD model of cognitive impairment
Treatment/Intervention	PO valsartan (10 mg/kg/d or 40 mg/kg/d), delivered in the drinking water for 5 mos
Results/Conclusion	Valsartan significantly promoted learning, acquisition and retention. It also significantly improved spatial, reference memory function during the probe test and resulted in a 2–3 fold reduction in soluble Aβ oligomers in the cerebral cortex, a reduction that coincided with their cognitive functional improvements. These beneficial effects were seen at doses ≤ those recommended for humans.

4.Telmisartan prevented cognitive decline partly due to PPAR- γ activation [124].
Animals	8 wk old adult male ddY mice given ICV Aβ 1–40
Model	Amyloid induced cognitive impairment
Treatment/Intervention	PO telmisartan or losartan (100 mg/kg/d) in drinking water (treatment duration 28d)
Results/Conclusion	Both telmisartan and losartan prevented Aβ 1–40 induced cognitive impairment. However the effect of losartan, which has a lower PPAR-ᵞ agonistic effect, was less than that of telmisartan. This may be partly due to PPAR-ᵞ activation (since co-treatment with to PPAR-ᵞ antagonist GW9662 partially inhibited this beneficial effect). Telmisartan also significantly reduced Aβ deposition, whereas losartan did not.

5.Angiotensin Receptor Blocker Prevented β-Amyloid–Induced Cognitive Impairment Associated With Recovery of Neurovascular Coupling [105].
Animals	8 wk old male WT, AD (APP23) and ICV Aβ injected mice
Model	Transgenic AD model and Amyloid induced cognitive impairment
Treatment/Intervention	Treatment started at age 8 wks and continued until age 13 wks. Experiment 1 APP23 and WT mice were treated with PO olmesartan (1 mg/kg/d) or vehicle. Experiment 2 Aβ ICV injected mice were treated with PO olmesartan (0.5 mg/kg/d, 1 mg/kg/d) or vehicle Experiments 3 and 4 Aβ ICV injected mice were treated with PO olmesartan (1 mg/kg/ d), hydralazine (30 mg/kg/d), nifedipine (10 mg/kg/d) or vehicle
Results/Conclusion	Olmesartan (0.5 and 1.0 mg/kg/d) significantly attenuated ICV Aβ induced cognitive dysfunction, independent of its Aβ or BP lowering effect. Hydralazine and nifedipine reduced BP like olmesartan but did not improve cognitive performance. In fact, they showed impairments comparable with that of vehicletreated animals. Olmesartan also significantly improved learning and memory in APP23 mice compared with vehicle treatment. It also significantly improved CBF, cerebrovascular reactivity and autoregulation and reduced ROS production to levels comparable with those of WT.

6.Candesartan improves memory decline in mice : Involvement of AT1 receptors in memory deficit induced by intracerebral streptozotocin [94].
Animals	8 wk old Swiss albino mice- ICV STZ (0.5 mg/kg/d) administered twice 48 h apart (days 1 and 3)
Model	STZ/Diabetes induced cognitive impairment
Treatment/Intervention	Early IP Candesartan sub-hypotensive doses (0.05 and 0.1 mg/kg/d) or vehicle days 1–14 Delayed IP Candesartan (0.1 mg/kg/d) started days 20–27
Results/Conclusion	Chronic candesartan treatment prevented STZ induced impairments in learning and memory. It significantly improved acquisition, retention and spatial memory. Such improvements were long lived when treatment began early but when treatment was delayed until substantial memory deficit was apparent, candesartan mediated improvement did not last. Candesartan also considerably reduced oxidative stress and free radical formation.

7.Telmisartan, a partial agonist of peroxisome proliferator-activated receptor γ , improves impairment of spatial memory and hippocampal apoptosis in rats treated with repeated cerebral ischemia [102].
Animals	250–300 g Male Wistar rats Day 1: vertebral arteries were electro-coagulated Day 2: Temporary BCCAO, using clips to interrupt CBF for 10 min followed reperfusion and repeated again 1 h later
Model	CCH induced cognitive impairment/VCI
Treatment/Intervention	PO telmisartan (0.3, 1, and 3mg/kg/d) was given for 7 d. On the 8th d, administration was performed either 1 h before (pre) occlusion and continued for another 6 d or administered 1 h (post) occlusion and continued for another 6 d (14 d treatment total)
Results/Conclusion	Chronic treatment with Telmisartan (pre- and post-ischemic) for 14 d significantly improved spatial memory and attenuated ischemia induced cognitive impairment in a dose-dependent manner. It also suppressed neuronal cell death (TUNELpositive cells) in the hippocampus after cerebral ischemia.

8.Nonhypotensive Dose of Telmisartan Attenuates Cognitive Impairment Partially Due to Peroxisome Proliferator-Activated Receptor- gamma Activation in Mice With Chronic Cerebral Hypoperfusion[99].
Animals	9 wk old male C57BL/6 mice- BCAS induced CCH
Model	CCH induced cognitive impairment/VCI
Treatment/Intervention	Low (non-hypotensive) dose: PO telmisartan (1 mg/kg/d) ± PO GW9662 (1 mg/kg/d) High (hypotensive) dose: PO telmisartan (10 mg/kg/d) ± PO GW9662 (1 mg/kg/d) or vehicle Treatment was initiated 7 days before BCAS surgery and continued until 30 days after BCAS
Results/Conclusion	Only the non-hypotensive dose of telmisartan ameliorated cognitive decline and improved spatial working memory in BCAS/CCH mice. It also significantly alleviated CCH induced microglial/astroglial activation, reduced cerebral mRNA expression of inflammatory cytokines (MCP-1 and TNF-α), reduced endothelial oxidative stress, oligodendrocyte loss and demyelinating changes in white matter. The protective effects against white matter lesions were, at least partially, mediated by PPAR-ɣ activation as they were partially offset by GW9662 cotreatment.

9.Blockade of AT1 Receptors Protects the Blood–Brain Barrier and Improves Cognition in Dahl SaltSensitive Hypertensive Rats [104].
Animals	5 wk old, male DSS/H rats (≈190 g)
Model	CCH induced cognitive impairment/VCI (hypertensive rats)
Treatment/Intervention	PO olmesartan at a nonhypotensive dose (1 mg/kg/d) for 4 weeks
Results/Conclusion	Olmesartan significantly ameliorated the cognitive deficits and markedly restored BBB function and expression of genes encoding occludin, claudin-5, collagen-IV, and MMP-9 without affecting BP in DSS/H rats.

10.Telmisartan protects against cognitive decline via up-regulation of brain-derived neurotrophic factor / tropomyosin-related kinase B in hippocampus of hypertensive rats [101].
Animals	12–14 wk old, SHRSPs
Model	CCH induced cognitive impairment/VCI (hypertensive rats)
Treatment/Intervention	Treated for 28 days with either: PO telmisartan (1 mg/kg/d) PO GW9662 (1 mg/kg/d)- PPAR-ɣ inhibitor PO telmisartan + PO GW9662 (1 mg/kg/d) PO telmisartan + TrkB antagonist ANA-12 (0.5 mg/kg/day) or PO vehicle
Results/Conclusion	Telmisartan treated SHRSPs showed significantly better spatial learning and reference memory than all other groups. Telmisartan protected against cognitive decline via up-regulation of BDNF/TrkB and partial activation of PPAR-ɣ in the hippocampus of SHRSPs, independent of BP-lowering.

11.Peroxisome Proliferator-Activated Receptor-gama Activation With Angiotensin II Type 1 Receptor Blockade Is Pivotal for the Prevention of Blood-Brain Barrier Impairment and Cognitive Decline in Type 2 Diabetic Mice [119].
Animals	15 wk old male KKAy (T2DM) and C57BL/6J mice
Model	Diabetes induced cognitive impairment
Treatment/Intervention	PO Telmisartan (1 mg/kg/d) or vehicle &/or PPARγ antagonist GW9662 (0.35 mg/kg/d) in drinking water for 7 wks
Results/Conclusion	KKAy mice treated with telmisartan had markedly improved cognitive function and preserved BBB integrity. Telmisartan also reduced swelling and MMP activity by its antioxidative and anti-inflammatory properties. This improvement was reduced with GW9662 Co-treatment.

12.Role of central angiotensin receptors in scopolamine-induced impairment in memory, cerebral blood flow, and cholinergic function [97].
Animals	12–15 wk old Swiss albino mice
Model	Anticholinergic (Scopolamine) induced cognitive impairment
Treatment/Intervention	PO Candesartan (0.05 and 0.1 mg/kg) was administered daily for 7 days and on the 7th day IP Scopolamine (3 mg/kg) was given
Results/Conclusion	Candesartan prevented scopolamine induced amnesia, restored CBF and ACh level, and decreased AChE activity and MDA level. However, the effect of the ARB candesartan on memory, CBF, ACh level, and oxidative stress was blunted by concomitant blockade of AT2R.The authors concluded that activation of AT1R appears to be involved in the scopolamine-induced amnesia and that AT2R contribute to the beneficial effects of candesartan.

13.Angiotensin II AT1 receptor blockers as treatments for inflammatory brain disorders [93].
Animals	9 wk old male C57BL/6 mice
Model	CCI (TBI) induced injury
Treatment/Intervention	SQ Candesartan (1 mg/kg/d)
Results/Conclusion	Candesartan treatment significantly improved spatial learning and memory, 4 wks after CCI injury. It was associated with a significant reduction in lesion volume, neuronal cell death and activated microglial cells. PPARɣ inhibition reduced, but didn’t eliminate the beneficial effect of candesartan.

14.Candesartan prevents impairment of recall caused by repeated stress in rats [90].
Animals	2 mo old Male Wistar rats (weight 140–160 g)
Model	Chronic stress induced cognitive impairment
Treatment/Intervention	PO Candesartan cilexetil (0.1 mg/kg/d) or vehicle for 21d
Results/Conclusion	Candesartan pretreatment effectively prevented chronic stress induced cognitive impairment, it significantly improved recognition memory as well as retrieval and active recall in chronically stressed animals

15.Effect of angiotensin II on spatial memory, cerebral blood flow , cholinergic neurotransmission , and brain derived neurotrophic factor in rats [89].
Animals	8–9 wk old male SD rats given ICV AngII (0.25 and 0.5 μg/side)
Model	AngII induced cognitive impairment
Treatment/Intervention	ICV Candesartan (0.5 and 1.0 μg/side) 5 min before ICV Ang II injection. Donepezil (5 mg/kg, PO) 1 h before ICV Ang II
Results/Conclusion	Ang II caused spatial memory impairment (affected acquisition, consolidation, and recall) with a significant reduction in CBF and ACh. Candesartan and donepezil both prevented Ang II-induced memory impairment, restored CBF and ACh levels.

16.Telmisartan attenuates cognitive impairment caused by chronic stress in rats [118].
Animals	Male Wistar rats, ≈150 g
Model	Chronic stress induced cognitive impairment
Treatment/Intervention	PO Telmisartan (1 mg/kg/d) or vehicle for 21d
Results/Conclusion	Telmisartan effectively restored cognitive function and significantly abolished the deleterious effects of chronic stress on recognition and long-term memory.

17.Angiotensin II type 1 receptor blocker losartan prevents and rescues cerebrovascular, neuropathological and cognitive deficits in an Alzheimer’s disease model [121].
Animals	~15 mo WT and AD (APP transgenic) mice
Model	Transgenic AD model of cognitive impairment
Treatment/Intervention	Prophylaxis - PO losartan (1 mg/kg/d) started at age ~2 month until age 8 months then increased to 10 mg/kg/d until 12mo (endpoint) Treatment - PO losartan (10 mg/kg/d) given for 3 mos, from age ~15mo until 18 mo (endpoint)
Results/Conclusion	Prophylaxis - Losartan completely prevented the onset of cognitive dysfunction, learning and memory deficits, in adult APP mice, despite unrelenting high levels of soluble Aβ species and Aβ plaque load Treatment - Losartan significantly improved memory acquisition, consolidation and recall performance, in aged APP mice without affecting Aβ accumulation/pathology.

18.The effects of valsartan on cognitive deficits induced by aluminum trichloride and D -galactose in mice [115].
Animals	2 mo old Swiss Albino Mice (25–30 g) given IP AlCl3 (10 mg/kg/day) and D-gal (150 mg/kg/day) for 90 days
Model	Experimentally induced sporadic dementia
Treatment/Intervention	IG Valsartan (20 mg/kg/day) for 60 days
Results/Conclusion	Valsartan prevented cognitive decline and improved learning and memory, restored cholinergic function and attenuated oxidative damage in AlCl3- and D-gal-treated mice.

19.Prophylactic angiotensin type 1 receptor antagonism confers neuroprotection in an aged rat model of postoperative cognitive dysfunction [96].
Animals	20 mo old male Sprague-Dawley rats
Model	Laporascopic surgery-induced cognitive decline in aged rats
Treatment/Intervention	IP candesartan (0.1 mg kg/d) for 14 days pre-treatment
Results/Conclusion	Chronic pre-surgical administration of low dose candesartan effectively prevented surgery-induced spatial learning and memory deficits. It also restored BBB function and reduced glial reactivity, NF-kB and neuroinflammation (IL-1β, TNF-α, and COX-2)

20.Cognitive enhancing effect of angiotensin-converting enzyme inhibitors and angiotensin receptor blockers on learning and memory [94]
Animals	Young (8–10 wk) and aged (35–38 wk) wistar rats given IP Scopolamine (1 mg/kg)
Model	Age and Anticholinergic (Scopolamine) induced cognitive impairment
Treatment/Intervention	PO ramipril (10 mg/kg/d 8d), IP perindopril (10 mg/kg/d 8d) IP losartan (20 mg/kg/d 8d), PO valsartan (20 mg/kg/d 8d), IP Piracetam (200 mg/kg/d 8d)
Results/Conclusion	Pretreatment with ACEIs and ARBs significantly improved the memory of aged rats and reversed scopolamine induced amnesia in both young and aged animals. ACEIs and ARBs were also superior to Piracetam (positive control) in improving cognition/memory.

21.Captopril and Valsartan May Improve Cognitive Function Through Potentiation of the Brain Antioxidant Defense System and Attenuation of Oxidative / Nitrosative Damage in STZ-Induced Dementia in Rat [109].
Animals	220–280g Adult male Wistar rats given ICV STZ (3mg/kg) at days 1&3.
Model	STZ/Diabetes induced cognitive impairment
Treatment/Intervention	PO captopril (50mg/kg/day) and valsartan (30mg/kg/day).
Results/Conclusion	Captopril and valsartan spared memory, reduced cognitive dysfunction and attenuated oxidative stress induced by STZ. These RAS modulators also seemed to potentiate superoxide dismutase (SOD) and catalase (CAT), antioxidant defense systems, in the brains of treated animals.

22.Angiotensin IV Receptors Mediate the Cognitive and Cerebrovascular Benefits of Losartan in a Mouse Model of Alzheimer’s Disease [122].
Animals	3-mo old male APP or WT mice
Model	Transgenic AD model of cognitive impairment
Treatment/Intervention	PO losartan (10 mg/kg/d) or vehicle for 1 mo then losartan animals were separated into: Cohort 1: ICV divalinal (AT4R blocker) or vehicle or in the final month of treatment
Results/Conclusion	Losartan-treated APP mice displayed significantly improved memory retrieval compared with untreated APP mice for all parameters. Losartan had no effect on memory performance in WT mice compared with WT controls. Divalinal countered losartan mediated benefits on spatial learning and memory. Losartan or divalinal administration does not alter Cortical and hippocampal Aβ pathology in App mice.

23.Chronic kidney disease accelerates cognitive impairment in a mouse model of Alzheimer’s disease , through angiotensin II [123].
Animals	9 mo old 5XFAD and wild-type mice fed an adenine-containing diet
Model	Transgenic AD model superimposed with CKD (accelerates cognitive impairment in AD mice)
Treatment/Intervention	PO Olmesartan (1 mg/kg/day) given once daily initiated 2 wks after the start of the adenine diet and continued for the remainder of the 10 wks
Results/Conclusion	Olmesartan significantly reduced the impairments in spatial learning and memory in addition to ameliorating BBB disruption and reducing hippocampal oxidative stress in 5XFAD with CKD.

24.Intranasal telmisartan ameliorates brain pathology in five familial Alzheimer ‘s disease mice [126].
Animals	2 month old 5XFAD mice
Model	Transgenic AD model of cognitive impairment
Treatment/Intervention	IN Telmisartan (1 mg/kg/day as 3 μL drop/nostril) or vehicle started at age 8 wks (continued for 5mos) 3 treatment groups as following: (i) Telmisartan -treated wild-type (WT) as the control group (ii) Telmisartan -treated 5XFAD mice (iii) vehicle-treated 5XFAD mice
Results/Conclusion	Telmisartan treatment improved hippocampus-dependent spatial acquisition and led to “goal-oriented performance” compared to vehicle treated animals. Telmisartan also ameliorated neuronal loss, amyloid burden and gliosis in the brains of 5XFAD mice, and caused a switch of microglia to the neuroprotective phenotype.

25.Candesartan , angiotensin II type 1 receptor blocker is able to relieve age- related cognitive impairment Pharmacological Reports Candesartan , angiotensin II type 1 receptor blocker is able to relieve age-related cognitive impairment [91].
Animals	Male young (2mo old) and Aged (24mo old) wistar rats
Model	Age associated cognitive decline
Treatment/Intervention	PO candesartan (0.1 mg kg/d) or vehicle for 21 days
Results/Conclusion	Candesartan significantly improved recognition memory in both young and aged animals. The degree of improvement in older rats was far more pronounced than in young rats. Candesartan significantly (P < 0.01) reversed age associated recall memory impairments and cognitive declines in aged rats.

26.Pleiotropic Benefits of the angiotensin receptor blocker candesartan in a mouse model of Alzheimer disease [92].
Animals:	3–4 mo old male and female C57BL6 APP mice
Model	Transgenic AD model of cognitive impairment
Treatment/Intervention	Cohort 1: SQ Candesartan (1 mg/kg/d) or vehicle for 2 mos Cohort 2: PO Candesartan (10 mg/kg/d) or vehicle for 5 mos
Results/Conclusion	Candesartan exerted potent anti-inflammatory effects, increased markers of neurogenesis & restored cerebrovascular reactivity, but unlike losartan had limited benefits on cognition. The authors suggest that PPAR-γ agonist properties are not needed for cognitive recovery with ARBs

27.Effects of enalapril and losartan alone and in combination with sodium valproate on seizures, memory, and cardiac changes in rats [116].
Animals	200–250g male wistar rats
Model	Sodium valproate induced cognitive impairment
Treatment/Intervention	IP enalapril (20 mg/kg/d), IP losartan (10 mg/kg/d) or vehicle
Results/Conclusion	Both ACEI (enalapril) and ARB (losartan) effectively prevented Sodium valproate induced cognitive impairment in seizure affected animals

28.A Comparative Study on the Memory-Enhancing Actions of Oral Renin-Angiotensin System Altering Drugs in Scopolamine-Treated Mice [113].
Animals	20–30g male Swiss mice given IP scopolamine
Model	Anticholinergic (scopolamine)-induced memory impairment
Treatment/Intervention	PO Captopril 25 mg/kg/d, Ramipril 4 mg/kg/d, Losartan 20 mg/kg/d, an hour before cognitive test
Results/Conclusion	All 3 RAS modulators showed a protective effect against scopolamine-induced memory impairment. These agents decreased the number of working and reference memory errors and spared short term and long term spatial memory.

29.Angiotensin Receptor Type 2 Activation Induces Neuroprotection and Neurogenesis After Traumatic Brain Injury [117].
Animals	9–10 wk old Mice
Model	TBI induced cognitive impairment
Treatment/Intervention	ICV infusion of CGP42112A (0.1, 1.0, or 10.0 ng/kg/min) or saline for 3d post-TBI
Results/Conclusion	CGP42112A attenuated TBI induced cognitive impairment (dose-dependent improvement in functional recovery& cognitive performance) accompanied by reduced lesion volume and induced neurogenesis

30.Direct stimulation of angiotensin II type 2 receptor enhances spatial memory [62].
Animals:	10–12 wk old male (WT, AT2R KO) C57BL/6 mice ICV Aβ (for 4 wks)
Model	Amyloid induced cognitive impairment
Treatment/Intervention	IP C21 (1, 3 or 10 mg/kg/d) for a duration of 2 wks administered with or without the bradykinin B2 receptor antagonist icatibant (70 mg/kg/d), IP infused via implanted osmotic mini-pump
Results/Conclusion	Treatment with C21 significantly enhanced cognitive function (spatial learning and memory) in WT, but not AT2RKO mice. This cognitive enhancement was attenuated by coadministration of icatibant. C21 also increased CBF in WT, but not AT2RKO mice. This increase was not observed in the icatibant co-treated group.

31.Direct angiotensin II type 2 receptor stimulation by compound 21 prevents vascular dementia [65].
Animals	10 wk old male C57BL6 (WT/ AT2KO) mice- BCAS induced CCH
Model	CCH induced cognitive impairment/VCI
Treatment/Intervention	IP C21 (0.01 mg/kg/d), vehicle or azilsartan (0.1 mg/kg/day) with or without PD123319 starting 1 wk before BCAS continued 6 wks
Results/Conclusion	C21and azilsartan treatments prevented BCAS induced cognitive impairment. These positive effects were attenuated with PD123319 and AT2KO in azilsartan treated and abolished in C21 treated animals. C21 also preserved CBF and reduced inflammatory cytokine expression with BCAS. The authors concluded that direct AT2R stimulation by C21 attenuates ischemic vascular dementia induced by hypoperfusion at least in part by an increase in CBF and reduced inflammation

32.RAS modulation prevents progressive cognitive impairment after experimental stroke: a randomized, blinded preclinical trial [60].
Animals	4 mo old male SHRs, t-MCAO induced cognitive impairment
Model	Post Stroke cognitive impairment (PSCI) in hypertensive animals
Treatment/Intervention	4 Groups: Sham, t-MCAO, IP C21(0.03 mg/kg/d 30d) t-MCAO, IP C21(0.03 mg/kg/d 7d) then IP candesartan (0.3 mg/kg/d) for the remainder of the study t-MCAO, IP saline 7d then IP candesartan (0.3 mg/ kg/d) for the remainder of the study
Results/Conclusion	Treatment with RAS modulators effectively preserved cognitive function, reduced cytotoxicity, and prevented chronic-reactive microgliosis in SHRs, poststroke. These protective effects were apparent even when treatment was delayed up to 7 days post-stroke and were independent of blood pressure and β-amyloid accumulation.

33.Angotensin receptor (AT2R) agonist C21 prevents cognitive decline after permanent stroke in aged animals-A randomized double- blind pre-clinical study [36].
Animals	14-mo old Male Wistar rats subjected to tandem d-MCAO
Model	Post Stroke cognitive impairment (PSCI) in aged animals
Treatment/Intervention	PO C21 (0.12 mg/kg/d 30d) or vehicle started 24h post ischemic insult and continued for 30d
Results/Conclusion	AT2R agonist, C21, effectively prevented development of PSCI and reduced cortical accumulation of Aβ1–42 in aged animals post-stroke

34.Delayed Administration of Angiotensin II Type 2 Receptor (AT2R) Agonist Compound 21 Prevents the Development of Post-stroke Cognitive Impairment in Diabetes Through the Modulation of Microglia Polarization [127].
Animals	12 wk old male Wistars +/− diabetes, subjected to t-MCAO
Model	Post Stroke cognitive impairment (PSCI) in diabetic animals
Treatment/Intervention	PO C21 (0.12 mg/kg/d) or vehicle started 3d post stroke for 8wks
Results/Conclusion	C21 resulted in a net cognitive improvement from baseline- 8 wks post- stroke in diabetic animals which may have been through modulation of the M1:M2 ratio.

35.Possible synergistic effect of direct angiotensin II type 2 receptor stimulation by compound 21 with memantine on prevention of cognitive decline in type 2 diabetic mice [129]
Animals	10-wk old KKAy mice
Model	Diabetes induced cognitive impairment
Treatment/Intervention	Four groups: (1) Control, (2) IP C21(10 μg/ kg/day), (3) PO Memantine (20mg/kg/day),(4) IP C21(10 μg/ kg/day) + PO Memantine (20mg/kg/day) for 4 wks
Results/Conclusion	Memantine and C21 had a synergistic beneficial effect in preventing diabetesinduced cognitive impairment. This synergistic effect may have occured through an increase in CBF and or ACh level.

36.Role of angiotensin system modulation on progression of cognitive impairment and brain MRI changes in aged hypertensive animals – A randomized double- blind pre-clinical study [34].
Animals	14-mo old male SHRs with UCCAO induced CCH, (aged SHRs)
Model	CCH induced cognitive impairment /VCI in aged hypertensive animals
Treatment/Intervention	PO candesartan (1 mg/kg/d 8wks), PO C21 (0.12 mg/kg/d 8wks) or vehicle started 24h post ischemic insult
Results/Conclusion	RAS modulators, Candesartan & C21 preserved cognitive function & prevented progression of VCI but only candesartan prevented loss of brain volume in aged hypertensive animals with chronic cerebral hypoperfusion.

Appendix D:

Table IV.

Clinical studies documenting the effect of renin angiotensin system modulation on cognition

1. Comparison of losartan and hydrochlorothiazide on cognitive function and quality of life in hypertensive patients [133].
Study design:	Double-blind randomized, control study
Patient population:	69 hypertensive patients ages 30 – 73 yo (mean 55 yo)
Study duration/ Follow-up:	A period of ~2.2 years (26 months)
Treatments:	Losartan (ARB), HCTZ (thiazide diuretic)
Study end-points:	Long-term changes in cognitive function and health state QoL index
Results/Conclusions:	Both drugs significantly lowered BP. Losartan improved health state QoL index in all subjects while HCTZ only improved it in patients ≥ 60 years old. Losartan also led to significant improvements in cognitive function, particularly memory, attention/concentration and comprehension while no changes in memory, attention/concentration or comprehension were observed in the HCTZ group. The authors concluded that losartan can have a positive effect not only on BP but also on impaired cognitive function, reversing even minimal cognitive deficits induced by hypertension

2. Influence of losartan and atenolol on memory function in very elderly hypertensive patients [130].
Study design:	Randomized double-blind active comparator parallel arm design
Patient population:	120 hypertensive patients mean age 81.3 yo
Study duration/ Follow-up:	A 6 months active treatment period (after 4-weeks wash-out)
Treatments:	Atenolol (βB) vs Losartan (ARB)
Study end-points:	Cognitive function (immediate memory, delayed recall, verbal fluency)
Results/Conclusions:	Although both atenolol and losartan had similar antihypertensive efficacy (no differences in BP lowering), only losartan significantly improved both immediate and delayed memory function

3. Effects of valsartan compared with enalapril on blood pressure and cognitive function in elderly patients with essential hypertension [110].
Study design:	PROBE active comparator parallel arm design
Patient population:	144 hypertensive patients age 61– 80 yo
Study duration/ Follow-up:	A 4 months active treatment period (after 2-weeks wash-out)
Treatments:	Enalapril (ACEI) vs Valsartan (ARB)
Study end-points:	Cognitive function (immediate memory, recognition, delayed recall, verbal fluency)
Results/Conclusions:	Both valsartan and enalapril had a clear antihypertensive effect, but the former led to a greater reduction at 16 weeks. Valsartan significantly improved both immediate and delayed memory function compared to both baseline and enalapril. Enalapril didn’t result in significant changes in any of the cognitive function tests

4. Use of angiotensin receptor blockers is associated with a lower incidence and progression of alzheimer’s disease matter lesions are present [139].
Study design:	prospective cohort analysis
Patient population:	Veterans aged ≥65 yo
Study duration/ Follow-up:	A 6-month period
Treatments:	ARBs, lisinopril (ACEI), other AH medications
Study end-points:	Incidence and progression of AD/ dementia
Results/Conclusions:	Patients taking ARBs exhibited a reduced incidence of AD/dementia compared to those taking lisinopril or other AHs. The use of ARBs was also associated with lower progression of AD/dementia compared to lisinopril group and other AHs. Therefore, ARBs are associated with a significant reduction in the incidence and progression of AD/ dementia compared to both ACEIs and AH medications.

5. Effects of hypertension therapy based on eprosartan on systolic arterial blood pressure and cognitive function: Primary results of the Observational Study on Cognitive function and Systolic Blood Pressure Reduction open-label study [136].
Study design:	Large, multinational open-label observational study
Patient population:	25, 745 newly diagnosed hypertensive patients mean age 65 yo
Study duration/ Follow-up:	A period of 6-months
Treatments:	Eprosartan 600 mg/d (ARB)
Study end-points:	Mean absolute change in SBP and cognitive/ MMSE score from baseline (relation between AH therapy with ARB and cognitive status)
Results/Conclusions:	Eprosartan based therapy was associated with overall improvements in cognitive function (MMSE score), which were related to the magnitude of BP reduction and support the use of ARB based AH treatment to delay or prevent cognitive decline hypertensive individuals

6. Use of angiotensin receptor blockers and risk of dementia in a predominantly male population: prospective cohort analysis [140].
Study design:	Prospective cohort analysis
Patient population:	819, 491 participants (98% male) aged ≥65 yo with CV disease
Study duration/ Follow-up:	A period of 5 years
Treatments:	3 cohorts (ARBs, ACEI-lisinopril, other CV drugs-βB, CCBs),
Study end-points:	Time to incident AD/ dementia, Disease progression (admission to nursing homes and death) among participants with pre-existing AD/ dementia
Results/Conclusions:	ARBs were associated with significant reductions in incident dementia compared with lisinopril or CV comparators. In patients with pre-existing AD/ dementia, ARBs showed significantly lower rates of admissions to nursing homes and death compared with users of the CV comparators Participants who used ARBs throughout the study or who started using ACEIs and switched to ARBs showed significantly lower risk of dementia than the group that used ACEIs throughout the study and did not switch to ARBs. The authors concluded that ARBs are associated with a significant reduction in the incidence and progression of AD and dementia compared with other AH agents.

7. Impact of Angiotensin Receptor Blockers on Alzheimer Disease Neuropathology in a Large Brain Autopsy Series [149].
Study design:	Multiple logistic regression analysis of patient data from the National Alzheimer Coordinating Center
Patient population:	890 hypertensive patients, mean age 81 yo
Study duration/ Follow-up:	Median time between enrollment and death was 2 years
Treatments:	ARBs, other AH meds, untreated
Study end-points:	Cognitive function (scores) and neuropathologic data (neuritic plaque, neurofibrillary tangle measures and vascular injury markers)
Results/Conclusions:	When comparing all 3 groups, those who received ARBs had significantly higher cognitive function and memory scores relative to the other 2 groups. Participants treated with ARBs were also less likely to have a clinical and neuropathologic diagnosis of AD. They showed less amyloid deposition, compared with both untreated participants and those treated with other AH medications including ACEIs.

8. Candesartan and cognitive decline in older patients with hypertension - A substudy of the SCOPE trial [134].
ClinicalTrials.gov Identifier	Foreign (UK)
Study design	Randomized double-blind, placebo-controlled clinical trial
Patient population	257 older adults (70–89 years) with hypertension
Study duration/ Follow-up	A period of 44 months
Treatments	8–16 mg candesartan or placebo once daily
Study end-points	Cognitive function
Results/Conclusions	The candesartan group showed significantly less decline in the cognitive domains of attention and episodic memory with a slight benefit in speed compared to placebo and no significant differences in working memory or executive function. The authors concluded that the potential for ARB–based antihypertensive therapy to reduce cognitive decline associated with hypertension in older adults may need further study.

9. Effects of telmisartan on cognition and regional cerebral blood flow in hypertensive patients with Alzheimer’s disease [148].
ClinicalTrials.gov Identifier	Foreign (Japan)
Study design	prospective randomized, open-label parallel design
Patient population	20 patients with hypertension and probable AD
Study duration/ Follow-up	A period of 6 months
Treatments	Telmisartan (40–80 mg/ day) or Amlodipine (5–10 mg daily)
Study end-points	Cognitive function, CBF, BP
Results/Conclusions	The telmisartan group showed similar or improved cognitive performance after 6 months, but the amlodipine group had worse performance on all tests after 6 months, although both groups had similar reductions in BP with telmisartan increasing CBF in multiple regions but amlodipine group increasing CBF only in the right cingulate gyrus. The authors concluded that telmisartan may have additional benefits making it useful for the treatment of elderly hypertensive patients with AD

10. Antihypertensive classes, cognitive decline and incidence of dementia: a network meta-analysis. [142].
Study design:	Meta-analysis / systematic review of RCTs
Patient population:	850,189 hypertensive patients with no prior cerebrovascular disease, mean age range 64–75yo
Study duration/Follow-up:	Median duration of 6 months
Treatments:	Diuretics, ARB, ACE-I, CCB, βB
Study end-points:	Cognitive function andincidence of dementia
Results/Conclusions:	AH treatment had significant benefits on overall cognition and may be effective for slowing cognitive decline and prevention of dementia with ARBs being most effective. ARBs were more effective than βB, diuretics and± ACEIs). The mean change in BP didn’t differ significantly between different AH drug classes.

11. Antihypertensive drugs decrease risk of Alzheimer disease- Ginkgo Evaluation of Memory Study [137].
Study design:	Secondary, post hoc, longitudinal analysis of GEMS trial (double-blind RCT)
Patient population:	older adults aged 75–96 yo with normal cognition (n = 5 1,928) or MCI (n = 5,320)
Study duration/ Follow-up:	6 years
Treatments:	Diuretics, ARB, ACE-I, CCB, βB
Study end-points:	Incidence of AD/ dementia among different AH medication user groups
Results/Conclusions:	Incidence of AD/ dementia among those with normal cognition and MCI at baseline was significantly lower in those using RAS modulators or diuretics. Authors concluded that diuretic, ARB, and ACE-I use was, in addition to and/or independently of BP, associated with reduced risk of AD/ dementia in participants with normal cognition, with similar trends among participants with MCI.

12. Renin-angiotensin system blockers affect cognitive decline and serum adipocytokines in Alzheimer’s disease. [146].
Study design:	Retrospective study
Patient population:	184 AD patients, age 76.4 ±7.9 years (range 52–90 years)
Study duration/ Follow-up:	The average length of follow-up was 2.45 ± 0.95 years (range 1–4.33 years)
Treatments:	ARB, other AH drugs, no AH drugs (in the case of normotensive patients)
Study end-points:	Cognitive function, Neuro-radiological assessment
Results/Conclusions:	The modified Fazekas scale scores for deep subcortical WMH and periventricular hyperintensities did not differ significantly among the 3 groups. Although the cognitive scores at onset/first visit were almost the same among all 3 groups, the annual decline in cognitive score was significantly lower in the ARB group compared to all other groups. The authors concluded that treatment with ARBs might slow the rate of cognitive decline in patients with AD and is a good therapeutic option with well-established safety for the management of hypertension in patients with AD.

13. The Antihypertensives and Vascular, Endothelial and Cognitive Function Trial (AVEC) [132].
ClinicalTrials.gov Identifier	NCT 00605072
Study design	Randomized Double-blind clinical trial
Patient population	53 Participants, age ≥ 60 years with hypertension and executive dysfunction
Study duration/ Follow-up	A period of 1 year
Treatments	Candesartan, Lisinopril, HCTZ
Study end-points	Cognitive function and cerebrovascular hemodynamics
Results/Conclusions	The candesartan group had the greatest improvement in executive function and cerebrovascular hemodynamics compared with both the lisinopril and HCTZ treatment groups. The authors concluded that in older adults with hypertension and mild executive dysfunction, an ARB-based regimen may be associated with preserved executive function, improved blood flow and cerebrovascular reserve compared to ACEI- or diuretic- based regimens, especially in those with relatively lower pretreatment blood flow velocity.

14. Effects of Centrally Acting Angiotensin Converting Enzyme Inhibitors on Functional Decline in Patients with Alzheimer’s Disease [155].
Study design	observational study- secondary analysis of the DARAD trial, a multicenter, blinded, randomized controlled trial
Patient population	406 patients with mild to moderate AD, age ≥ 50 years
Study duration/ Follow-up	A period of 12-months
Treatments	Central ACEIs (Ramipril, perindopril, lisinopril, trandolapril and fosinopril) vs other BP meds
Study end-pts	Cognitive and memory function, psychiatric
Results/Conclusions	There 12-month rate of cognitive decline was a significantly lower, 25% difference in patients taking central ACE-Is, compared to the other BP meds. This remained significant after adjusting for age, gender, education, and BP. The authors concluded that central ACEIs are associated with a reduced rate of functional decline in patients with AD, without an association with mood or behavior, suggesting that these agents may slow AD progression.

15. Associations of Anti-Hypertensive Treatments with Alzheimer’s Disease, Vascular Dementia, and Other Dementias [29].
Study design:	Prospectively recorded, nested case-control study
Patient population:	9,197 aged ≥ 60 yo diagnosed with dementia
Study duration/ Follow-up:	Duration ≥ 6 months
Treatments:	ARB, ACEI, other AH drugs
Study end-points:	Associations of ARBs and ACE-Is with VaD/AD/unspecified other dementia
Results/Conclusions:	Patients prescribed either ARBs or ACE-Is were less likely to develop VaD, AD and unspecified/other dementia (inverse associations) than patients prescribed other AH drugs. Inverse associations were strongest for ARBs compared with ACE-Is. There were inverse dose-response relationships between ARBs and ACE-Is with AD.

16. Antihypertensive drugs, prevention of cognitive decline and dementia: A systematic review of observational studies, randomized controlled trials and meta-analyses, with discussion of potential mechanisms [143].
Study design:	Systematic review of observational studies, RCTs and Meta-Analyses
Patient population:	Study populations consisted of 1,346,176 subjects, mean age 74 years
Study duration/ Follow-up:	Range 6 months-9 years
Treatments:	AH medications
Study end-points:	Incidence and progression of cognitive decline/dementia
Results/Conclusions:	AH therapy, especially ARBs, ACEIs and CCBs, could reduce the incidence and progression of cognitive impairment/dementia, not only by lowering BP but also through other inherent neuroprotective properties.

17. Antihypertensive drug use and risk of cognitive decline in the very old: an observational study-The Newcastle 85+ Study [135].
ClinicalTrials.gov Identifier	Foreign (U.K.)
Study design	population-based observational cohort study
Patient population	238 older adults aged 75–96 yo with normal cognition (n=5 1,928) or MCI (n = 5,320)
Study duration/ Follow-up	A period of 3 years
Treatments	Diuretics, ARB, ACE-I, CCB, βB
Study end-points	Rate of cognitive decline over a 3 yr period
Results/Conclusions	Unadjusted analysis for AH use and cognitive change over 3 years suggested less cognitive decline was associated with ARBs and CCBs only. These remained significant only for CCBs after adjustment for confounders. On the other hand, βBs were associated with a significantly higher rate of cognitive decline. No significant associations were found for ACE-I or diuretics

18. Renin-Angiotensin-System Modulation may slow the Convension from Mild Cognitive Impairment to Alzheimer’s Disease [150].
Study design	longitudinal
Patient population	2,520 participants with MCI, age ≥ 75 years
Study duration/ Follow-up	A maximum follow-up period of 5 years
Treatments	RAS medication group (ARB, ACE-I), non-RAS medication group (Diuretics, CCB, βB)
Study end-pts	Cognitive, functional decline and AD conversion rate
Results/Conclusions	Even though BP control was statistically better among non-RAS medication users compared to RAS medication users, RAS users were a lot less likely to convert to AD. RAS users also demonstrated slower declines in memory and attention than non-RAS users. This slowing of cognitive decline was even more pronounced with BBB-crossing RAS medications, which were also associated with better executive function, language and global cognition compared to all other groups. Atenolol didn’t induce changes in cognitive performance The comparison between losartan and atenolol was significant for both memory functions. The authors concluded that in very elderly hypertensive patients, chronic AT1 receptor blockade by losartan could improve cognitive function

19. Associations of centrally acting ACE inhibitors with cognitive decline and survival in Alzheimer’s disease [147].
ClinicalTrials.gov Identifier	Foreign (U.K.)
Study design	retrospective observational study
Patient population	5,260 patients receiving acetylcholinesterase inhibitors who use Centrally acting-ACEIs (C-ACEIs); Noncentrally acting ACEIs (NC-ACEIs) or neither, at the time of AD diagnosis.
Study duration/ Follow-up	A period of 9 months
Treatments	Centrally acting-ACEIs l; Noncentrally acting NC-ACEIs
Study end-pts	Change in cognitive function as measured by MMSE
Results/Conclusions	MMSE scores significantly increased by in patients on C-ACEIs and deteriorated by in those on NC-ACEIs, with no differences in survival. The authors concluded that in people with AD, already receiving AchE inhibitors, those also taking C-ACEIs had stronger initial improvement in cognitive function, but there was no evidence of longer-lasting influence on dementia progression.

20. Memory is preserved in older adults taking AT1 receptor blockers [138].
Study design	A longitudinal study
Patient population	1,626 participants without dementia, age 55–91 years
Study duration/ Follow-up	A period of 3 years
Treatments	ARBs, other AH meds (Diuretics, CCB, βB, ACEIs, α1-blockers, α2-agonists, or direct vasodilators)
Study end-pts	Cognition, MRI brain volume and WMH
Results/Conclusions	The non-ARB group performed worse than normotensives on all measures of cognition, verbal learning, immediate and delayed recall, over the 3-year follow-up, while ARB users did not differ from normotensive subjects on any measure of cognition and demonstrated better recognition memory than those taking other antihypertensive medications. These cognitive differences are especially notable, given that the ARB group had significantly more participants diagnosed with type 2 diabetes, which has been associated with a 1.5- to 2.5-fold greater risk of dementia. Even with this added risk factor, the ARB users demonstrated preserved memory function over 3 years of follow-up. In short, hypertensive participants demonstrated worse memory and executive function, as well as greater memory decline, over the 3-year follow-up than normotensives, unless they were ARB users, who showed preserved memory compared with those taking other antihypertensive drugs.

21. Pharmacogenetics of Angiotensin-Converting Enzyme Inhibitors in Patients with Alzheimer’s Disease Dementia [141].
ClinicalTrials.gov Identifier	Foreign (Brazil)
Study design	Prospective pharmacogenetics study
Patient population	Late onset AD patients mean age 65 yo
Study duration/ Follow-up	A period of 1 year
Treatments	ACEIs vs other meds
Study end-points:	Cognitive function
Results/Conclusions:	No functional impacts were found regarding any genotypes or pharmacological treatment. Either for carriers of ACE haplotypes, or for APOε4- carriers, ACEIs slowed cognitive decline independently of BP variations. APOε4+ carriers were not responsive to treatment with ACEIs. ACEIs may slow cognitive decline for patients with AD, more remarkably for APOε4- carriers of specific ACE genotypes

22. The association of multiple anti-hypertensive medication classes with Alzheimer’s disease incidence across sex, race, and ethnicity[145].
Study design	A retrospective cohort study
Patient population	1,343,334 users of six different AH drug treatments, age ≥ 67 years
Study duration/ Follow-up	Data from 2007–2013
Treatments	ARBs, other AH meds (Diuretics, CCB, βB, ACEIs)
Study end-pts	Cognition, MRI brain volume and WMH
Results/Conclusions	Results: RAS-acting AH meds were more protective against AD than nonRAS-acting ones for males. ARBs were superior to ACEIs for both white men and white and black women.The authors concluded that ARBs may, reduce AD risk, particularly for white and black women and white men.

23. Observational Study of Brain Atrophy and Cognitive Decline Comparing a Sample of Community-Dwelling People Taking Angiotensin Converting Enzyme Inhibitors and Angiotensin [152].
ClinicalTrials.gov Identifier	Foreign (Australia)
Phase	II and III
Study design:	A longitudinal study
Patient population:	565 participants with T2D, HTN or both age 55–90 years without dementia
Study duration/ Follow-up:	3.2 years
Treatments:	ARBs, ACEIs, other AH, no AH
Study end-pts:	Cognition, MRI brain volume
Results/Conclusions:	Neither ACEI nor ARB use was associated with cognitive decline. Patients taking an ARB had a slower rate of brain atrophy than those taking an ACEI, independent of BP control.

24. Exploiting Drug-Apolipoprotein ε Gene Interactions in Hypertension to Preserve Cognitive Function: The 3-City Cohort Study [153].
ClinicalTrials.gov Identifier	Foreign (Australia, France)
Study design	A Prospective multisite, population-based cohort study
Patient population	3359 persons using antihypertensive drugs (median age 74 yo)
Study duration/ Follow-up	10 years follow-up
Treatments	Centrally acting ACEI, (captopril, fosinopril, lisinopril, perindopril, rampril, and trandolapril), peripherally acting ACEIs (benazepril, enalapril, moexipril, and quinapril) ARB, βb, CCB and thiazide-like drugs
Study end-pts	Cognitive function, assessed at baseline, 2, 4, 7 and 10 years using a validated battery of test covering global cognition, verbal fluency, immediate visual recognition memory, processing speed and executive function.
Results/Conclusions	This study showed that exposure of APOε4 carriers to ACEIs or ARBs over time was associated with better general cognitive function, compared with other antihypertensive drugs. Findings did not support RASs’ lipophilicity or ability to cross the BBB as a potential mechanism. The authors concluded that the use of RAS blockers for hypertension in APOε4 carriers may improve long-term cognitive function in older populations at risk of cognitive decline and dementia

Appendix E:

Table V-.

Clinical trials in progress to study the effect of RAS modulators on cognition

1. Telmisartan vs. Perindopril in Hypertensive Mild-Moderate Alzheimer’s Disease Patients (SARTAN-AD) [80].
ClinicalTrials.gov Identifier:	NCT02085265
Condition or disease	Alzheimer’s Disease and Hypertension
Study design	Randomized, Open Label Clinical Trial with Parallel drug Assignment
Estimated Enrollment	240 participants
Intervention/treatment	- Telmisartan 40 mg or 80 mg/day (depending on age and tolerability) - Perindopril 2 mg, 4 mg or 8 mg/day (depending on renal function and tolerability)
Phase	Phase II
Study Start Date	March 2014
Estimated Study Completion Date	March 2021
Primary Outcome Measures	BP, Ventricular enlargement , Safety, Adverse events
Secondary Outcome Measures	Change in hippocampal volume, volume of grey and white matter, measures of cognitive function
Eligibility Criteria	Age ≥ 50 Years
Inclusion criteria	Probable or possible AD dementia Established diagnosis of hypertension
Exclusion criteria	Intolerance or contraindications to study medications FAD form of Alzheimer’s disease

2. Health Evaluation in African Americans Using RAS Therapy (HEART) [81].
ClinicalTrials.gov Identifier:	NCT02471833
Condition or disease	Alzheimer’s Disease
Study design	Randomized Double blinded, Interventional Clinical Trial
Estimated Enrollment :	66 participants
Primary Purpose:	Prevention
Intervention/Treatment	Telmisartan 20mg or Telmisartan 40mg or Placebo each given orally once a day for 8 months
Phase	Phase I
Study Start Date	April 2015
Estimated Completion Date	June 30, 2020
Primary Outcome Measures	Change in concentration of CSF angiotensin metabolites [ From baseline to 8 months]
Secondary Outcome Measures	Changes in plasma RAS components (Renin, ACE, aldosterone) Change in CSF T-tau, P-tau Cognitive function (MMSE, WAIS, MoCA, Set-shifting Test, Flanker Inhibitory Control and Attention Test)
Eligibility Criteria	Age ≥30 Years
Inclusion criteria	African American with hypertension and Family history of AD
Exclusion criteria	Mean resting SBP ≥110 and ≤ 170 mmHg Current use of RAS acting medication History of stroke, Dementia, Heart failure or Diabetes Types I and II

3. Candesartan’s Effects on Alzheimer’s disease And Related Biomarkers (CEDAR)[158].
ClinicalTrials.gov Identifier:	NCT02646982
Condition or disease	Mild Cognitive Impairment
Study design	Randomized Quadruple Blinded (Participant, Care Provider, Investigator, Outcomes Assessor)Interventional Clinical Trial
Estimated Enrollment :	72 participants
Intervention/Treatment	Candesartan vs matched Pacebo once daily for 12 months Candesartan will be started at 8 mg orally, once daily. The dose will be increased in 2 week increments to 16 mg and 32 mg once daily, as tolerated (SBP>110 mm Hg, DBP>40 mm Hg with no reported symptoms of hypotension).
Phase	Phase II / Phase III
Study Start Date	June 2016
Estimated Completion Date	September 2021
Primary Outcome Measures	Changes in the following: number of subjects with hypotension, number of hypotensive episodes, Scr and serum potassium
Secondary Outcome Measures	Changes in CSF phospho-tau, Aβ42, cytokines Change in arterial stiffness and cerebral vasoreactivity Changes in levels of circulating EPCs Changes in cognitive function as assessed by (ADAS-cog, CDR, EXAMINER, Spatial 1-Back test) Change in WMH assessed by MRI
Eligibility Criteria	60 – 85 Years (Adult, Older Adult)
Inclusion criteria	Mild Cognitive Impairment, with abnormal memory function and delayed recall (MoCA < 26) but general functional performance sufficiently preserved and Aβ positivity determined
Exclusion criteria	Intolerance to ARBs Current use of antihypertensive medication including ARBs or ACEIs Renal disease or uncontrolled congestive heart failure Inability to have MRI History of increased intracranial pressure or bleeding diathesis Women of childbearing potential (non-menopausal)

4. Risk Reduction for Alzheimer’s Disease (rrAD) [157].
ClinicalTrials.gov Identifier:	NCT02913664
Condition or disease	Cognitively Normal Older Adults
Study Type	Interventional (Clinical Trial), Randomized, Factorial Assignment
Estimated Enrollment :	640 participants, 60 Years to 85 Years (Adult, Older Adult), all sexes.
Intervention/Treatment	Drug: Angiotensin II receptor blocker (ARB, losartan) and calcium channel blocker (amlodipine)
Phase	Phase II
Study Start Date	September 2016
Estimated Completion Date	September 2022
Primary Outcome Measures	Change in global neurocognitive function [ Time Frame: 2 Years ]
Secondary Outcome Measures	Assessment of domain-specific neurocognitive function Assessment of whole brain and hippocampal volume Assessment of global and regional brain perfusion Assessment of brain white matter hyperintensity (WMH) Assessment of brain white matter microstructural integrity
Eligibility / Inclusion criteria	Age 60–85, all races/ethnicities, and both sexes are eligible. A positive family history of dementia defined as having at least one first-degree relative with a history of AD or other type of dementia or b) having subjective cognitive decline.
Exclusion criteria	Clinically documented history of stroke, focal neurological signs or other major cerebrovascular diseases Diagnosis of significant neurologic diseases Uncontrolled diabetes mellitus Allergy to angiotensin receptor blockers (ARBs)