Genistein,A Phytoestrogen,Delays the Transition to Dementia in Prodromal Alzheimer’s Disease Patients

INTRODUCTION

As the global population ages, the prevalence of Alzheimer’s disease (AD) continues to rise, posing significant social, economic, and healthcare burdens worldwide. Despite decades of research, effective treatments for AD remain elusive, highlighting the complex nature of this neurodegenerative disorder.

One crucial aspect that has emerged in recent years is the importance of early detection and intervention in AD management [1]. Early diagnosis allows for timely implementation of therapeutic strategies, potentially slowing disease progression and improving patient outcomes. However, diagnosing AD in its early stages remains a considerable challenge, as symptoms often manifest subtly and are easily overlooked or attributed to normal aging processes. Thus, there is an urgent need for the development and validation of reliable biomarkers and diagnostic tools to facilitate early detection and intervention [2]. For instance, it has been suggested that deficits detected in recall, familiarity, or false recognition in older people could be used as early prodromal markers of cognitive impairment [3].

Moreover, until recently the current therapeutic landscape for AD primarily consists mainly of symptomatic treatments that offer modest symptomatic relief but do not modify the underlying disease pathology [4]. To effectively combat AD, there is a critical imperative to develop disease-modifying treatments that target the fundamental mechanisms driving neurodegeneration. Recently, monoclonal antibody therapies like aducanumab or lecanemab hold the promise of halting or even reversing disease progression, thereby offering hope for significantly altering the course of AD and improving long-term outcomes for affected individuals [5].

Furthermore, accumulating evidence suggests that AD is a multifactorial disease, influenced by a complex interplay of genetic, environmental, and lifestyle factors. Traditional approaches targeting single pathological pathways have shown limited efficacy, underscoring the need for comprehensive, multifaceted treatment strategies that address the diverse etiological factors contributing to AD pathogenesis.

In this paper, we pay particular attention to the importance of early detection, and the recognition of AD as a multifactorial disorder that needs a multifactorial treatment.

We report that genistein is able to delay the transition of prodromal AD to dementia and that it acts via multiple synergistic mechanisms.

OXIDATIVE AND REDUCTIVE STRESS IN ALZHEIMER’S DISEASE

The pioneering work of George Perry and Mark Smith shed light on the presence of oxidative stress in the brains of AD patients [6]. Their research revealed that oxidative stress is a consistent feature across all analyzed AD patients and could elucidate a significant portion of the disease’s pathological manifestations.

In our investigations, we examined oxidative stress in both AD patients and experimental disease models, particularly the APP/PS1 mouse model [7]. Recognizing the criticality of probing disease manifestations in its early stages, we explored oxidative stress in individuals at risk of AD, specifically those with the APOE 4/4 risk genotype. Surprisingly, we observed reductive stress in these at-risk individuals, despite the absence of evident mild cognitive impairment [8].

Systemic oxidative stress was quantified through parameters such as lipoperoxidation or glutathione oxidation in mononuclear cells. We interpret the emergence of reductive stress as a cellular hyper-response to initial oxidative stress triggered by interactions between amyloid-β (Aβ), proteins, and mitochondria. This interaction likely disrupts normal mitochondrial function by binding to cytochrome heme groups, thereby escalating oxidative stress. Subsequently, cellular defenses activate antioxidant mechanisms, resulting in an overshooting of these reactions leading to reductive stress (Fig. 1).

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Fig. 1

Reactive oxygen species (ROS) mediate the link between amyloid-β (Aβ) and tau hyperphosphorylation via p38MAPK.

Recently, our longitudinal study spanning a decade has revealed the eventual dissipation of reductive stress, coinciding with the onset of oxidative stress and early signs of neuronal damage, heralding prodromal AD and its progression to dementia [9].

LIFESTYLE MODIFICATIONS FOR ALZHEIMER’S DISEASE PREVENTION AND TREATMENT

The profound impact of AD on individuals and society has spurred extensive research aimed at treating or delaying dementia onset. While efforts such as antibody production against Aβ or tau proteins have garnered attention, we advocate for the pivotal role of lifestyle modifications, particularly in preventing dementia progression among individuals with prodromal AD and mild cognitive impairment [10].

Evidence from our research and others underscores the efficacy of physical exercise in delaying dementia onset in prodromal AD patients and ameliorating cognitive decline in animal models like the APP/PS1 mice [11].

Neurostimulation, dietary alterations, and selective supplementation offer additional avenues with demonstrated benefits in slowing dementia progression and minimal side effects. Notably, lifestyle changes have been shown to significantly impact disease progression, instilling hope for patients to potentially delay the transition to dementia [10].

ACTIVATING RXR/PPARγ AS A THERAPEUTIC APPROACH TO TARGET AMYLOID-β

Since Alois Alzheimer’s time, the significance of amyloid plaques in disease development has been recognized [12]. Subsequent elucidation of Aβ protein and its production mechanisms has spurred endeavors to attenuate disease progression by reducing Aβ levels and impeding plaque formation.

Pioneering work by Gary Landreth demonstrated the therapeutic potential of bexarotene, a cancer therapy drug, in AD treatment [13]. Bexarotene’s activation of retinoid X receptor (RXR) and peroxisome proliferator-activated receptor gamma (PPARγ) facilitates apolipoprotein E (ApoE) production, promoting Aβ clearance from the brain and subsequent degradation (Fig. 2). For a review, see [14]. Our research corroborates the favorable impact of bexarotene in experimental AD treatment.

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Fig. 2

Anti-amyloid-β action of genistein or bexarotene is mediated by activation of the PPARγ/RXR receptor driving ApoE production that drags amyloid-β into the blood plasma for subsequent degradation in the liver.

While monoclonal antibodies targeting Aβ have also been explored, outcomes have varied, with some showing clinical improvement while others demonstrate inconclusive results. Recent developments such as FDA approved lecanemab and aducanumab show promise, yet optimization through further research remains imperative to enhance efficacy.

GENISTEIN IN EXPERIMENTAL ALZHEIMER’S DISEASE TREATMENT

Our interest in genistein stems from previous investigations into the longevity disparity between sexes (in many species like Wistar rats and humans), wherein estrogens were found to induce longevity genes, including antioxidant genes [15, 16]. However, concerns over estrogen administration limitations prompted exploration of phytoestrogens like genistein, which preferentially binds to the beta-estrogen receptor without feminizing effects or heightened cancer risks.

Moreover, genistein’s interaction with PPARγ receptors presented a rationale for evaluating its therapeutic potential in AD (Fig. 2). Thus, we conducted experiments in the APP/PS1 mouse model to assess genistein’s potential benefits in mitigating AD pathology.

The advantage of genistein over bexarotene and, in general, over most treatments proposed as anti-amyloid therapies, which are monoclonal antibodies, is that genistein can be administered orally and presents no serious side effects. Some reports in the media without any scientific basis have cited the possibility that genistein may cause cancer. There is no evidence in the worldwide literature to support this claim, instead, the current literature supports an anticancer effect of genistein [17–21]. Naturally, no increase in the incidence of cancer has been observed in countries with high intakes of genistein in the diet, such as Japan and other countries in East Asia.

The effect of genistein is mediated by the activation of RXR/PPARγ, which increases the production of ApoE, which binds to the amyloid protein and drags it out of the brain into the blood, to be eventually metabolized in the liver. Figure 3 shows the effects of genistein administration in AD mice, analyzing two parameters that are highly important as signs of the disease: cognition measured by spatial orientation (Hebb-William’s test) and the ability to distinguish odors [22]. The figure shows values for healthy animals (black bar), animals suffering from the disease (red bar), and animals that have taken genistein (blue bar), bexarotene (orange bar), or both (green bar). It can be observed in Fig. 3 that we have been able to reproduce the results of bexarotene, which protects transgenic mice against the toxicity of Aβ peptide, although the effects of genistein are equal or superior to those of bexarotene and naturally do not have the undesirable side effects of the latter. Therefore, we believe that genistein is superior to bexarotene for treating experimental AD [23]. Bagheri et al., using different model of AD, showed that pretreatment with high doses of genistein improves cognition in Aβ injected rats [24].

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Fig. 3

Genistein improves cognition and odor recognition in APP/PS1 mice.

Naturally, finding a treatment for a disease in an experimental model only serves as a basis for a solid trial in humans, which always involves clinical trials, which we outline below.

GENISTEIN DELAYS THE TRANSITION TO DEMENTIA IN PRODROMAL ALZHEIMER’S DISEASE PATIENTS

The next step in our work consisted of proposing a clinical trial (NCI clinical trial NCT01982578) to determine if genistein is useful for delaying the transition to dementia in patients who present the early signs of AD, known as prodromal AD patients.

The primary objective of the trial was to analyze Aβ deposition in the brains of prodromal AD patients who were administered genistein or a placebo for up to 12 months. Aβ deposition in the brain was assessed using positron emission tomography with flutemetamol as the tracer.

The secondary objective of this trial was to analyze changes in cognitive tests (see Table 1) to assess if genistein administration delayed cognitive decline measured by these tests.

The cognitive tests utilized in the study included the Mini-Mental State Examination, the Memory Alteration Test, the Clock Drawing Test, the Complutense Verbal Learning Test (TAVEC), the Delayed TAVEC Test, the Revised Barcelona Test for Semantic Fluency, the Revised Barcelona Test for Phonological Fluency, the Rey Complex Figure Test, the Rey Test measured in percentages, the Delayed Rey Test, and the Delayed Rey Test measured in percentages. Table 1 indicates the cognitive characteristics measured by these tests.

Figure 4 illustrates that in all cases, patients treated with genistein experienced less cognitive decline than those treated with a placebo. This reached statistical significance for individual tests in the case of the direct TAVEC and the Rey Test measured in percentages [25].

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Fig. 4

Genistein treated Prodromal AD patients lose less cognition than placebo-treated patients in a 12-month evolution period. The meaning of each of the tests indicated is in Table 1.

Furthermore, the fact that in all tests, patients taking genistein experienced less cognitive loss than those taking a placebo (although in many cases the individual difference was not significant, probably due to the small number of cases), led us to seek an integrated test that captured changes occurring in all the aforementioned tests. A similar approach was used by Mintun et al. when studying the effect of donanemab in AD [26]. These researchers used a composite score and observed that donanemab was superior to placebo in delaying cognitive decline in patients with early AD.

Our combined test involved studying the differences between the end and beginning of treatment for each patient treated with genistein or a placebo. The composite AD score was derived through a two-step process: firstly, by computing the discrepancy between the assorted scores at the twelve-month interval and their respective baseline values, and subsequently, by aggregating all variables. Consequently, lower composite scores denote a deteriorated cognitive condition. Within our cohort, the composite score ranged from –156.2 to 11. Analysis employing this metric revealed that patients receiving genistein treatment exhibited significantly superior performance compared to those administered placebos (genistein-treated patients displayed a mean score of –14.83 (SD = 24.18), whereas placebo recipients scored –55.89 (SD = 53.28); resulting in a mean difference of 41.06 (95% CI: 5.13 to –76.99), p = 0.028). This observation may be attributable to the consistently superior performance of genistein-treated patients across all administered tests, although statistical significance was only observed in two specific tests [25].

Finally, we report the results of the primary objective, which is Aβ deposition in the brain. Back in 2012, a study reported that genistein was able to reduce Aβ positive aggregates in the lateral blade of the dentate gyrus in a rat model of AD using immunohistochemistry techniques [27]. To our knowledge, we are the firsts to provide data from humans. Our determinations were made by positron emission tomography (PET) using flutemetamol, and we observed that in the anterior cingulate gyrus, flutemetamol deposition was lower in patients treated with genistein compared to those treated with a placebo, meaning that deposition increased in patients treated with placebo but not in those treated with genistein [25]. Recent lipidomic research has shown that the cingulate gyrus is one of the brain areas most affected by aging and plays a fundamental role in both cognition and emotions [28]. Therefore, the results of Aβ deposition coincide with the analysis of cognitive tests, showing that genistein is a treatment that may delay the transition to dementia in patients with prodromal AD.

THE MULTIMODAL MECHANISM OF ACTION OF GENISTEIN

When it comes to intervening in a multifaceted disease like AD, it is always interesting to approach treatment from various mechanisms of action. The idea that a drug must act through a single mechanism of action to interfere with a disease has been surpassed. This is one of the issues we encounter in the treatment of AD with monoclonal antibodies: they are highly specific to Aβ protein, acting solely through one mechanism of action, namely reducing Aβ load.

Figure 5, modified from our previous work [29], shows that the action of genistein is at least tetra-modal, meaning it acts through four different mechanisms. Firstly, we described that it acts by activating antioxidant genes [30]. It does not have (as previously believed) a significant antioxidant effect due to its chemical properties, but rather acts much more effectively by promoting synthesis through gene regulation mechanisms of antioxidant enzymes such as glutathione peroxidase, Mn-superoxide dismutase, or catalase.

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Fig. 5

The multimodal action of genistein as an antioxidant, anti-inflammatory, anti-amyloid-β (Aβ) and antiapoptotic agent (taken from Mas-Bargues et al., [29]).

Furthermore, genistein exhibits an anti-inflammatory role, which is well observed in experimental studies where neuroinflammation in the brains of APP/PS1 mice treated with genistein is analyzed through staining for Iba 1. The figure shows variations in interleukins that determine the predominance of the anti-inflammatory state in the presence of genistein [31].

Genistein also serves as an anti-amyloid therapeutic (which is the primary focus of this work) as it promotes the production of ApoE, which is responsible for capturing Aβ and removing it from the brain into general circulation, all mediated by its interaction with the PPARγ receptor.

Finally, genistein plays a crucial role in autophagy, especially mediated by the activation of lysosome biogenesis and the increase in their activity [32]. We should note that the increase in autophagy mediated by genistein helps eliminate aberrant proteins along with other altered molecules which, if accumulated, could induce programmed cell death, i.e., apoptosis. In this sense, treatment with genistein can be understood to act as an anti-apoptotic treatment [29].

CONCLUSION

Lifestyle interventions are important in the prevention of the transition to AD of patients with minimal cognitive impairment. We recently reviewed evidence of the importance of physical exercise in the prevention and treatment of AD [11]. Here we report evidence that administration of genistein, a phytoestrogen found in many foods, especially soya, rapidly improves cognition and Aβ deposition in experimental AD. Moreover, in a double-blind placebo-controlled bicentric pilot clinical trial it significantly delays the transition to dementia in prodromal AD patients. The magnitude of change is similar to that reported in monoclonal antibody-based treatments. These results underpin the importance of food supplement administration in the management of early AD.

AUTHOR CONTRIBUTIONS

Cristina Mas-Bargues (Writing – original draft; Writing – review & editing); Jose Vina (Conceptualization; Funding acquisition; Visualization; Writing – original draft; Writing – review & editing); Consuelo Borrás (Conceptualization; Funding acquisition; Writing – original draft; Writing – review & editing).