Ginkgo biloba

7 Ginkgo biloba

From Traditional Medicine to Molecular Biology

Yves Christen Ipsen

Paris, France

I. INTRODUCTION

A botanical curiosity, Ginkgo biloba is also extremely original in its bio- chemical composition. This probably explains the multiplicity of effects observed with Ginkgo biloba extract—EGb 761. Studied at all levels of the organization of life (molecular, cellular, tissular, the entire organism, and behavior, including in humans), EGb 761 has shown particularly interesting effects in four domains: protection of the nervous system, protection of the circulatory system, protection against various diseases of the retina, and protection against some otorhinolaryngeal (ORL) diseases. Several of these effects are explained by its antioxidant action and by its effects on gene expression. As far as physiology and therapy are concerned, one remarkable aspect of EGb 761 is that it does not have a single unidirectional effect (activator or inhibitor) but rather is regulatory, promoting the adaptation of the organism to the situation at hand. This result may be due to the fact that it is a natural extract, that is, a set of molecules made by the process of natural

F IGURE 1 Number of publications on Ginkgo biloba published every year. Incomplete for the year 2001. Data Christen from the ginkgo database. (Courtesy of F. Colmant, Medipsen, Paris.)

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selection, a mechanism specialized in the making of adaptive biological systems.

Before any of its potential therapeutic uses were known, Ginkgo biloba was the object of study by botanists because it is an extremely original tree. For more than 30 years, an extract from the tree has also been a very popular drug, first in Europe (especially in France and Germany) and, more recently, in the United States. While several extracts of Ginkgo biloba exist, the reference extract, the extract that exists in standardized forms and has been the object of most of the published studies, is that known as EGb 761 (extrait

de Ginkgo biloba nj761). In recent years, the number of publications about Ginkgo biloba has multiplied impressively. I have described the universe of ‘‘ginkgology’’ elsewhere as an expanding universe (1). The concept of ginkgology is justified by the existence of a significant collection of data in very different domains but all concerning Ginkgo biloba: botany, evolution, ethnology and the study of traditional cultures, cultural anthropology, Asian civilizations and Far Eastern mythology, chemistry, pharmacology, medicine, etc. From this point of view, Ginkgo biloba is singular, perhaps unequaled in the medical world.

The increase in the number of scientific publications related to ginkgo confirms the reality of this expansion, as the analyses contained in the ginkgo database (Fig. 1) show. Although some of these studies concern nonmedical aspects of ginkgology, most examine EGb 761 and its interest to medicine. This situation is a rather paradoxical phenomenon in the history of the med- ical and pharmaceutical sciences: in most cases, drugs are studied primarily before their sale is approved and immediately afterward. The demands of marketing have led pharmaceutical laboratories to neglect them later, because they think they have said all that there is to be said about them and because they have found and are focusing on the next generation of drugs. Histori- cally, the singularity of this difference is even greater than would appear from

a simple reading of the statistics about the number of publications. Today, the important drugs arrive on the market with their history already established. The sponsors must argue convincingly that the research that led to their development was perfectly planned and rational and led to the specific target defined at the outset (e.g., inhibition of an enzyme or binding to a specific receptor). We might say that these drugs, conceptually, are represented in a finished universe (even though it is always possible to add new indications). EGb 761, on the other hand, exists in an open universe, into which totally new and even unforeseen data can arrive.

II. WHAT IS GINKGO BILOBA? Ginkgo biloba is a tall, long-lived tree originating from China (2). Ginkgo

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of the leaves. This species, which was classified in 1771 by Linne´, is the only representative of the class of Ginkgoatae and even of its entire phylum, that of Ginkgophyta, distinct from all other trees, notably Coniferophyta and Cycadophyta. To this taxonomic singularity is added its unique status as a living fossil. Ginkgo trees already flourished 180 million years ago, when several species existed. Over the past million years, Ginkgos have progres- sively disappeared from most of the earth, finally surviving only in China. They have in fact nearly disappeared there, except for several rare areas, the best known of which are in Zhejiang province on the west peak of Tianmu Mountain (3). Fortunately, this disappearance in the wild has been compen- sated for by the cultivation of numerous specimens in China, Japan, other countries in the Far East, and even throughout the rest of the world. Many of these protected trees are very old. There are reports of more than 100 specimens in China that are more than 1000 years old (4). Ginkgo biloba was first introduced in Europe in 1690 by the botanist Engelbert Kaempfer, who described it as a ‘‘tree with duck feet’’ owing to the shape of its leaves (5). It is also an extremely hardy tree, as proved by its ability to survive the atomic bomb on Hiroshima. Today it adjusts well to pollution and is resistant to parasites; both reasons account for its success in our cities.

III. WHAT IS GINKGO BILOBA EXTRACT? Ginkgo biloba extract, EGb 761, is a standardized extract of Ginkgo biloba. It

contains 24% flavonoids and 6% terpene lactones. The flavonoids are nearly exclusively flavonol-O-glycosides (combination of the phenolic aglycones— quercetin, kaempferol, or isorhamnetin—with glucose or rhamnose or both in different positions of the flavonol moiety). The terpenoids are ginkgolides (3.1% of the extract) and bilobalide (2.9%), both found exclusively in the Ginkgo biloba tree. Ginkgolides are diterpenes, five of which have been identified (gingkolides A, B, C, M, and J). Bilobalide is a sesquiterpene. EGb 761 also contains proanthocyanidins (prodelphinidins) and organic acids (6). The combined activity of all the components of EGb 761 is required for its optimal therapeutic action (6). Several other extracts exist, especially in United States, where this herbal drug is not strictly regulated as in Europe. These extracts can vary in content, purity, and potency.

IV. CLINICAL INDICATIONS The use of the ginkgo tree in traditional Chinese medicine is very ancient. The

seeds have been used since at least the thirteenth century and the leaves since at least the sixteenth century. Modern Chinese pharmacopoeias mention the use of the leaves in the treatment of cardiac and pulmonary diseases but also

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extract is much more recent and results from research inspired not by traditional medicine but linked instead to the development of pharmacology.

A first extract was marketed in 1965 in Germany by Dr. Willmar Schwabe Company. EGb 761 was first registered in France in 1974 and sold in 1975 by Institut Ipsen (now Ipsen Pharma) under the trademark Tanakan and in Germany as Rokan by Intersan in 1978 and as Tebonin forte by Dr. Willmar Schwabe Company in 1982. Its first therapeutic indications involved circula- tory disturbances. More recently, the therapeutic effects that have been studied most are those affecting the psychobehavioral disorders associated with aging. These include cognitive disturbances, such as age-related memory disorders (grouped together today as mild cognitive impairment, MCI), or dementia, such as Alzheimer’s disease. Whether these various indications (Alzheimer’s disease, in particular), are included in the approved indications varies from country to country. EGb 761 is also prescribed against several diseases of the sensory organs that are often associated with aging, such as presbyacusis, tinnitus, age-related macular degeneration, glaucoma, etc.

In Europe (principally in France and Germany) these indications are officially approved, and patients are reimbursed for these health expenses. In the United States, on the other hand, Ginkgo biloba extract is mostly con- sidered an herbal drug and is marketed as a health food.

In addition to the indications cited, EGb 761 has been the subject of numerous investigations that have reached definitive conclusions about its action in extremely diverse areas. Some of these indications are justified by serious studies—placebo-controlled and double-blinded—while others rest on much more fragile or uncontrolled foundations. Among the former, we note cognitive impairments associated with multiple sclerosis (8); mood disorders, particularly depression (9); sexual dysfunctions (10); and radia- tion-related disorders (11).

V. BIOLOGICAL PROOFS OF EFFICACY Numerous studies have reported the principal effects of EGb 761: against

neurodegenerative disorders, visual or ORL diseases, the ischemic process, and the vascular diseases associated with it (Table 1). These studies concern all levels of the organization of life: molecular, cellular, tissular, the entire organism (especially when aged or diseased), behavioral, and finally human. It is this multiplicity of findings that in its entirety constitutes the best proof of the extract therapeutic effect (12).

Many of these effects are ubiquitous. Thus, EGb 761 protects the cellular organelles, mitochondria in particular (13), the membranes, DNA, etc., and the cell as a whole against stress of diverse origins (radiation, toxic

150 T ABLE 1 Main Effects of Ginkgo biloba Extract EGb 761 and Its Clinical Applications

Clinical trial and Therapeutic field

Molecular studies and

epidemiological study Memory impairment

mechanisms of action

Animal models

Dual coding test (51); EEG and MCI

Prevention: protection

Many experiments in rats and

against oxidative stress

mice (49); protection of

study (52) and beneficial

(27); immediate effect: on

hippoccampus in old

effects on memory (53),

attention and vigilance (54) Alzheimer’s disease

gene expression (32,48)

animals (50)

Protection against oxidative

Beneficial effect in

Symptomatic effect in

stress (55) and against

transgenic mice (59);

multicenter study (61);

agregation and toxicity of

enhancing of synaptic

preventive effect in

beta-amyloid (25,56);

plasticity in the perforant

epidemiological stucy

enhancing of alpha-

pathway (60)

(Epidos; 16)

secretase activity (57); induction of heme- oxygenase1 (33,58)

Multiple sclerosis

Beneficial effect on

Beneficial effect on attention,

experimental allergic

memory, and functioning in

encephalomyelitis in rat

patients (8)

Parkinson’s disease

Protection of aged

Protection against the

Some positive data in

mitochondria (13);

neurotoxic effects of

uncontrolled observations

induction of Hsp70 (63)

MPTP (64,65)

Amyotrophic lateral

Protection against oxidative

Protective effect in the

transgenic mice expressing

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the SOD1 mutation (66)

Stroke

Protection against oxidative

Protection against cerebral

Beneficial effect in patients

stress

ischemia in gerbils and

with cerebrovascular with cerebrovascular

insufficiency; beneficial Ginkgo

effect after cerebral infarct Age-related macular

lesions

Beneficial effect in patients degeneration

Protection against free

Protection of the retina in old

radicals (67) and against

rats (69); protection

biloba

loss of gluthatione (68)

against light-induced retinal degeneration (70)

Diabetic retinopathy

Protection against free

Protection against retinal

Beneficial effect in patients

radicals (27)

injuries in diabetic rat (72);

effect on intraocular proliferation (73)

Glaucoma

Protection against free

Effect on circulation to the

Increasing of ocular blood flow

velocity in patients (77) Vertigo

radicals (75)

optic nerve (76)

Beneficial effect on

Beneficial effect in patients

vestibular compensation in

cat, rat, and guinea pig (78–80)

Hearing loss and

Beneficial effect in patients sudden deafness

Improving survival of primary

Effect on models of cochlear

auditory neurons after

pathologies associated

trauma (Pujol and Puel,

with ischemia (82)

personal communication)

Tinnitus

Attenuation of salicylate-

Beneficial effect in patient

induced tinnitus (84)

Peripheral arterial

Several placebo-controlled occlusive disease

Enhancing of the release of

Vascular protection;

vasodilating factors such

vasodilator effect (37)

multicenter studies;

as NO/EDRF (37)

beneficial effect on intermittent claudication (87,88)

Trophic disorders and

Beneficial effect on leg ulcers vasomotor changes

Hemorheological and

(90); beneficial effect in 151 in the extremities

metabolic effects (89)

Raynaud’s syndrome (91)

T ABLE 1 Continued

Clinical trial and Therapeutic field

Molecular studies and

epidemiological study Premenstrual

mechanisms of action

Animal models

Effective against the syndrome

Vasoregulatory properties

congestive symptoms, particularly breast symptoms and neuropsychological symptoms (92)

Altitude sickness

Anti-ischemic, antihypoxic,

Beneficial effect during a

vasoregulatory, metabolic,

Himalayan expedition (93)

and antiedema properties (89)

Protection against

Inhibition of the clastogenic ionizing radiation

Protection against free

radicals; inhibition of

factor among the Chernobyl

liquidators (11) Depression

clastogenic factor (11)

Regulatory effect on

Beneficial effect on several

Beneficial effect on mood

neurotransmitters

models of stress

among depressed patients

Sexual dysfunction

Effect on phosphodiesterase

Vasodilator effect (89)

Beneficial effect on sexual

dysfunction (10) Christen

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of aging and not only neurodegenerative diseases. This explains the diversity of the indications for EGb 761.

To these ubiquitous effects we can add some that are much more specific, revealed by numerous studies, especially in vivo, in various pathophysiolog- ical models. For example, EGb 761 acts in vivo in models of various nervous system diseases: Alzheimer’s disease (transgenic mice overexpressing a histo- pathological mutation of amyloid precursor protein), amyotrophic lateral sclerosis (transgenic mice expressing a harmful mutation to the superoxide dismutase gene, SOD1), Parkinson’s disease (animals exposed to MPTP), stroke, head trauma, etc. EGb 761 also acts against cardiovascular disease, affecting various models of arrhythmia, myocardial ischemia and hypoxia, cerebral edema, etc. (12,14,15).

Finally, several clinical studies have confirmed its effect in the corresponding human diseases. These studies, as well as experimental re- search, justify expansive, large-scale prospective studies to determine whether EGb 761, which is associated with a lower prevalence of Alzheimer’s disease in the Epidos epidemiological study (16), can prevent or delay the onset of this disease. The ginkgo evaluation of memory (GEM) study in United States (17) and the GuidAge study in France (18) are currently among the largest preventive studies in this field.

The clearest demonstration of the effects of EGb 761 does not concern a disease in the strict sense of the term but rather involves the duration of life. EGb 761 is the only substance that has been shown to be able to increase mean and especially maximum longevity in rats (19), which means a species of mam- mals. This longevity effect has also been found in the worm Caenorhabditis elegans (20). Although this effect does not actually involve protection against

a disease, its medical importance is beyond dispute: it unambiguously demonstrates the reality of the biological action and proves how well this substance is tolerated.

VI. MECHANISMS OF ACTION All of the various effects of EGb 761 cannot be attributed to a single mech-

anism of action or to a single molecule in the extract. The free-radical scav- enging effect, for example, is due to flavonoids (21) but also to ginkgolides (22), the PAF-inhibiting effect (23) as well as the inhibition of peripheral benzodiazepine receptors (24) to ginkgolides, the protection against apoptosis and beta-amyloid toxicity to flavonoids (25), the increased longevity, at least in the worm C. elegans, to flavonoid action (20), and the upregulation of the cytochrome c oxidase gene to the biobalide (26).

Reduction to a single mechanism is certainly not necessary. Nonetheless we can explain many of EGb 761’s effects by its antioxidant action and its

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experimental data have confirmed that it acts as a free-radical scavenger (21,27), including in vivo in humans (22). This finding tends to explain its protective role—of the nervous system in particular—in the harmful processes associated with aging and with traumatic, ischemic, or toxic injuries: most of the neurodegenerative disorders seems to be linked to a similar mechanism, i.e., the aggregation of toxic proteins and the effect of free radicals (28,29). More recently, effects have been observed on gene expression: EGb 761 in- hibits AP-1 transcription factor (30) and increases or decreases the expression of many genes in vitro (31). Studies have also shown that EGb 761 affects in vivo gene expression in animals (32). In several cases, the gene effect involves not only mRNA expression but also production of the corresponding protein, including proteins with a clear-cut protective role, especially against neuro- degenerative diseases, such as heme oxygenase 1 (33).

The free-radical-scavenging action is linked to the effect on gene ex- pression, in that many genes, including those for transcription factors such as AP-1 and NF-kappaB, are sensitive to the free-radical concentration in cells. Nonetheless, from a biological and treatment point of view, these two mechanisms must be distinguished. Antioxidant action, in the strict sense of the term, explains the protection against degenerative mechanisms and thus the prevention or slowing down of decay. EGb 761 also exerts—for example, on attention mechanisms—effects that occur immediately or at least rapidly and cannot be explained simply by protection against cell death. Some of these effects may be due to changes in gene expression, which is a fast process. This particular mechanism can bring into play a multitude of targets within the cell, since proteins are often expressed in clusters within cells. The idea of a single target is thus illusory. The multiplicity of effects is particularly expected here because the extract contains numerous distinct molecules. The tools of molecular biology now offer new means of approaching this complexity, since DNA arrays make it possible to see, in a single experiment, an effect on several thousand genes. These are thus particularly important methods for testing the effect of such complex treatments as plant extracts, especially EGb 761 (34).

Another protective mechanism involved in many physiological and pathological situations is protection against cell death, especially apoptosis (25,35,36). Such an effect is potentially beneficial in many neurodegenerative diseases. Interestingly, EGb 761 protects cells from apoptosis in aged animals but it also exerts a cytostatic effect on tumor cells, thus showing an interesting adaptive effect (see latter on).

VII. AN ADAPTIVE EFFECT One of the most remarkable aspects of EGb 761 is that, unlike many modern

drugs, its pharmacological effects do not simply inhibit or activate a given

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sometimes antagonistic. Thus, at the vascular level, it does not simply exert effects on arteries, veins, and capillaries: these effects can be dilatational or constrictive, depending on the target or condition (6,37).

We know other examples of EGb 761’s regulatory effects, in particular its action on apoptosis. Many experiments have shown that it protects against apoptotic phenomena, in particular in the framework of neurodegenerative processes (25), where this type of cell death plays a harmful role (38). In the presence of tumor development, however, EGb 761 acts in the opposite direction; that is, it has cytostatic properties that promote the elimination of cancers (39).

The effect of EGb 761 on the cerebral metabolism exemplifies another of these adaptive mechanisms. EGb 761 promotes oxygen and glucose con- sumption in the brains of animals in ischemic situations (40), but, inversely, facilitates metabolic economy in healthy subjects (41). PET imaging studies in humans have shown that the most gifted subjects tend to expend only a limited amount of brain energy for a given task compared with other individuals (42), and, inversely, that individuals with dementia use more energy than normal subjects (43). Economical energy metabolism is thus essential for optimal brain operations.

From a behavioral and neurochemical point of view, the effect of EGb 761 is similarly regulatory, promoting appropriate decision-making and neurotransmitter concentrations that help restore normality (44).

This drug thus seems to exert an adaptive effect; that is, it is sensitive to the physiological or pathological situation in which the organism finds itself (45). This may explain why it increases longevity. We note in this regard that the effects of EGb 761, which seem to differ from those of other drugs, resem- ble to some extent those of calorie restriction. The latter also extend life span and protect neurons against cerebral aging (46), undoubtedly by general ho- meostatic adaptation of the organism. By regulating energy expenditures (and modifying gene expression), calorie restriction preserves metabolic potential and limits free-radical production, itself associated with oxygen metabolism. The comparison is nonetheless incomplete, since EGb 761 intake is not accompanied by a reduction in the size of the organism, but it looks prom- ising, especially because calorie restriction could be not very well adapted to human condition. Recent data suggest that biomarkers of calorie restriction may predict longevity in humans even in people who were not really on a diet (47). That means that it could be physiologically possible to mimic the effect of calorie restriction. EGb 761 could influence the metabolism in this way.

VIII. BOTTOM-UP VERSUS TOP-DOWN EFFECT Modern therapeutic research is essentially based on the study of mechanisms

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cifically against them. The efficacy of the products thus developed should be guaranteed by the logic of this approach: they should work because they act directly on one of the steps involved in pathogenesis. This is what we might call

a bottom-up strategy, which involves starting from an elementary target, located downstream, to exert a systemic effect upstream. The efficacy of this strategy requires no further proof We nonetheless note two aspects that raise questions about this type of approach. The first is that, in reality, it is impossible to define any single target in a cell that is independent from all other elementary targets. Developments in cellular and molecular biology show that cell proteins function in clusters. They have a social life: by touching one, we affect many others. Along the same line, we note that all cell signaling systems constitute cascades that involve many more or less ubiquitous enzymes. The concept of a single, well-defined target is thus less simple than we imagine, and a product ‘‘designed’’ to affect only one enzyme or receptor will, by a sort of domino effect, affect a multitude. To this rather theoretical comment we must add another, more bothersome in practice: the highly targeted strategies often present side effects that are the logical consequence of damage to a natural physiological system that has a rationale. In some way, disease is always a steady state or a state of compensation, in which the organism is readapting to cope with a situation—very imperfectly, of course— which is why the organism is sick. The dislocation of one of the cogs of this system is thus also a disturbance, the disruption of a compensation.

This bottom-up strategy can conceptually be opposed by another, which we can describe as top-down. Rather than aiming at a simple target, we focus on a physiological whole, hoping to modify specific molecular targets by a cascade effect. Psychotherapy is an example of this approach. It does not target a particular receptor or neuromediator (as psychoactive drugs do) but the subjects themselves. When it works, this strategy must also necessarily affect molecular targets but without focusing on them. Other examples of top-down strategies include more technologically sophisticated practices of modern science, such as transcranial magnetic stimulation or the more invasive stimulation used in the treatment of Parkinson’s disease: a global target is aimed at, and the effect descends in a cascade to the molecular level.

Both strategies—bottom-up and top-down—have been used in EGb 761 studies. While many investigations have been based on a highly molecular strategy (for example, studies of the expression of some well-defined genes), others are more comprehensive (for example, its antistress effect or its effect on the duration of life, which is probably the most global parameter imag- inable). The pursuit of this twofold line of research is incontestably original. The hope is that the two approaches may, as often as possible, meet at some level. This is apparently the case in neurodegenerative diseases, where we

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observe a comprehensive protective effect that may be explained by the data available from the study of elementary mechanisms. Other situations seem less clear; for example, some nearly immediate cognitive effects are difficult to explain by neuroprotection, as is the regulatory action of EGb 761 (that is, it works so often in the direction of adaptation to circumstances). We might simply postulate that this particularly beneficial aspect is related to the fact that EGb 761 is a natural extract, shaped by natural selection for millions of years to interact with physiological targets and to adapt to cellular complex- ity. There is hardly any reason for a manufactured synthetic molecule essentially designed to interact with a single target to have such a charac- teristics. From this point of view, it is hardly surprising that, by acting on its target, this molecule disturbs the rest of the physiological system. A natural extract (i.e., shaped by natural selection) has a greater chance of being less disruptive.

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