Citicoline and CMT: Citicoline To Treat Motor Neuron + Demyelinating Diseases

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Citicoline and CMT:

CITICOLINE TO TREAT MOTOR NEURON DISEASES AND DEMYELINATING DISEASES

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2F26633 & DISPLAY=DESC

(Note: this is a long article, but it does mention Charcot Marie

Tooth disease)

The present invention relates to a method of treating motor neuron

and demyelinating diseases, including amyotrophic lateral sclerosis

(ALS) and multiple sclerosis (MS). In particular, the invention

relates to the use of citicoline (cytidine-5'- diphosphocholine or

CDP-choline) in the treatment of these diseases.

Background Of The Invention: There are a large number of

identifiable motor neuron diseases and demyelinating diseases. The

most common diseases of these types are ALS and MS, respectively.

ALS, also known as Lou Gehrig's disease, is a progressive disease of

the nervous system marked by muscular weakness and atrophy with

spasticity and hyper-reflexia due to degeneration of motor neurons

of the spinal cord, medulla and cortex of the brain.

Motor neurons are classified as either myelinated or non-myelinated

and their membranes are comprised of mainly neuronal lipids,

sphingomyelin, phosphatidylcholine, phosphatidylethanolamine, and

phosphatidylserine, each of which may contain varying amounts of

acyl phospholipids. ALS is characterized by a progressive loss of

upper and lower motor neurons, which includes the loss of the lipid

membranes.

Motor neurons are among the largest of all nerve cells in the brain

and spinal cord, and function to send messages to muscles throughout

the body. In ALS, motor neurons die and the muscles do not receive

these messages. As a result, muscles weaken as they lose their

ability to move. Eventually, most muscle action is affected,

including that which controls swallowing and breathing, as well as

the movement of major muscles in the arms, legs, back and neck.

There is, however, no loss of sensory nerves, so patients with ALS

retain their sense of feeling, sight, hearing, smell and taste.

Their mental capacity also remains relatively undiminished, and

hence ALS patients remain fully cognizant. While the course of ALS

is extremely variable and it is difficult to predict the rate of

disease progression, the majority of patients with ALS progressively

weaken and die over a three-to-five year period.

The first signs of ALS are often arm and leg weakness, muscle

wasting and faint muscle rippling.

These symptoms occur because muscles are no longer receiving the

nutrient signals they need for growth and maintenance--a result of

motor neurons dying. ALS nerve degeneration may also cause muscle

cramps and vague pains, or problems with speech and swallowing.

ALS occurs in two forms. In hereditary or familial ALS, a defective

gene is passed to successive generations. This accounts for about

10% of reported cases. The remaining 90% of cases are of unknown

etiology.

Recent research on inherited ALS has led to the discovery of a

defective gene believed to affect an enzyme called superoxide

dismutase. This enzyme rids the body of free radicals, which, if not

eliminated, can cause nerve cells to die. Free radicals are also

associated with a number of other diseases, and are even implicated

in the aging process itself.

Recently available treatments for ALS include medications which

increase survival time and/or aid quality of life by maintaining

muscle function.

Examples of such medications include various nerve growth factors.

While these treatments represent an incremental therapeutic advance,

a cure for ALS remains elusive. Thus, the primary treatment of ALS

still focuses on managing symptoms with physical, occupational,

speech, respiratory and nutritional therapy. For instance, drugs and

the application of heat or whirlpool therapy may help to relieve

muscle cramping. Moderate exercise can help maintain muscle strength

and function.

Drugs have also been used to treat fatigue, but may worsen muscle

cramps. GDNF, or Glial Cell-Derived Neurotrophic factor, has been

investigated in ALS, and is administered directly into the brain.

The drug had been shown to promote the growth of nerve cells in

animals. Pain in the limbs is common, persistent and hard to

control. Nonsteroidal anti-inflammatory drugs and simple analgesics

may bring relief. When pain is a major problem, opiates are used in

doses that are effective in relieving the pain.

Spasticity is a relatively minor problem which is caused by a lower

motor neuron lesion reducing the number of functioning motor units

to such a degree that the muscles are more hypo-than hypertonic.

Baclofen is the drug of choice for muscle spasms. Riluzole and is

often also used to treat ALS.

Other motor neuron disorders exhibit symptoms similar to ALS but may

have one or more recognizable differences. Kennedy's disease (X-

linked spinobulbar muscular atrophy) is an X-linked lower motor

neuron disease characterized by progressive muscular atrophy usually

beginning in mid-adult life. This disease is distinguished from ALS

by the absence of hyperreflexia and spasticity.

Adult Tay Sach's disease is caused by Hexosaminidase enzyme

mutations that produce lower motor neuronopathies that closely mimic

ALS. The disease is slowly progressive and may include dysarthria

and cerebellar atrophy. Spasticity may also be present, but is rare.

Spinal muscular atrophy (SMA), is a group of familial disorders

which affect large lower motor neurons. Muscle tissue often exhibits

evidence of denervation atrophy. Infantile SMA (SMA I, Werdnig-

Hoffman disease) is rapidly fatal, death generally ensuing within

the first year of life. Chronic childhood SMA (SMA II) progresses

slowly, beginning in childhood. Juvenille SMA (SMA III, Wohlfart-

Kugelberg- Welander disease) generally has a late childhood or early

adolescence onset and runs a slow course.

Primary lateral sclerosis (PLS) is rare disorder arising

sporadically in adults from mid-to late-life.

Symptoms include progressive spastic weakness of the limbs with

spastic dysarthria and dysphagia.

Fasciculations, amyotrophy and sensory changes are absent.

Familial spastic paraplegia (FSP) is a hereditary disease

characterized by progressive spastic weakness which begins in the

distal lower extremities.

Progressive neural muscular atrophy is a collection of degenerative

disorders characterized by progressive weakness and wasting of

skeletal muscles combined with sensory changes. The most common

example is Charcot-Marie-Tooth (CMT) disease. This and many other

progressive neuromuscular atrophy diseases are hereditary.

Various syndromes of progressive visual loss may also be attributed

to neurodegenerative disorders.

Examples include various forms of Friedreich's ataxia which are

characterized by a slowing of conduction in the optic nerves. There

are two broad categories of visual pathology, namely, selective

degeneration of retinal ganglion cells with secondary optic atrophy

and a more diffuse degeneration involving all retinal components. An

example of the latter is retinitis pigmentosa.

Of the various demyelinating diseases, MS is by far the most well

known. The causative agent of MS is unknown, although both

infectious agents and autoimmunity are suspected. MS often strikes

in early adulthood and is characterized by the formation of lesions

(demyelinated plaques) in the central nervous system. The afflicted

individual can exhibit lack of coordination, dysarthria (slurred

speech), numbness, paralysis and/or urinary incontinence. Blindness

has also been reported. Unlike the linear progression of ALS,

however, the course of MS often involves spontaneous remission

followed by relapses.

Fortunately, in a majority of cases, permanent remission eventually

occurs, but only after successively less severe relapses, and with

an average duration of illness spanning 27 years.

Despite the fact that many MS patients experience eventual

remission, the long life span of the disease inflicts psychological

and economic hardship on patient and caregiver alike. Hence,

treatment of at least the symptoms of the disease is highly

advantageous.

Another demyelinating disease is acute disseminated

encephalomyelitis (ADEM). This disease is distinguished from MS by

having a monophasic course.

It is often associated with immunization (postvaccinal

encephalomyelitis) or infection (postinfectious encephalomyelitis).

Symptoms include widely scattered small foci of perivenular

inflammation and demyelination and may be chronic in nature.

Acute hemorrhagic leukoencephalitis (AHL) is characterized by

perivenous demyelination and intense infiltration by mononuclear and

polymorphonuclear inflammatory cells. The clinical course resembles

severe forms of ADEM, but may be even more explosive in onset and

progression. Death may occur within two to four days of onset,

although complete neurologic recovery has been observed.

Occasionally, as with ADEM, the disease may take a chronic course

similar to MS.

Although there are some treatments for these diseases, there remains

a need in the art for a method of treating motor neuron and

demyelinating diseases, such as ALS and MS, which provides for both

increased relief of symptoms and at least temporary cessation or

even reversal of neuronal damage.

Summary Of The Invention The present invention meets this need by

providing a method for treating motor neuron and demyelinating

diseases which comprises administering an effective amount of

citicoline or a pharmaceutically-acceptable salt thereof. The

present invention further provides a method and composition for

treating motor neuron and demyelinating diseases which comprises

administering an effective amount of a combination of citicoline and

a glucocorticoid, preferably dexamethasone, prednisone or

methylprednisolone. The present invention also relates to the use of

citicoline for the preparation of a pharmaceutical medicament for

the treating of motor neuron and demyelinating diseases, comprising

admixing an effective amount of citicoline with a pharmaceutically

acceptable carrier.

It is, therefore, one object of the invention to provide methods for

improving the treatment of symptoms of those afflicted with a motor

neuron or demyelinating disease.

Another object of the invention is to provide methods for decreasing

symptoms in patients who have suffered nerve injury or nerve death

due to motor neuron or demyelinating disease.

Still another object of the invention is to provide methods for

preventing the worsening of symptoms over the course of the disease,

i. e., to inhibit progression.

These and other objects of the invention will be apparent to those

of ordinary skill in view of the discussion above and the additional

detailed description provided below relating to preferred

embodiments of the invention.

Detailed Description Of The Preferred Embodiments Citicoline is

believed to have multiple therapeutic effects. Although the relative

contribution of each effect on the treatment of motor neuron or

demyelinating disease is unknown, citicoline and its metabolites--

which include cytidine and choline--are believed to play important

roles in the generation of phospholipids involved in membrane

formation and repair. These compounds also are believed to

contribute to critical metabolic functions, such as the formation of

nucleic acids, and the synthesis of the neurotransmitter

acetylcholine. Thus, under conditions of frank neuronal damage with

associated nerve cell degeneration, citicoline may function to: (1)

stabilize membranes by providing substrates for membrane

maintenance; (2) repair damaged membranes by supplying important

substrates for membrane formation; and, (3) restore neuronal

function by supplying a substrate for the formation of

acetylcholine. Moreover, unlike other proposed therapeutic agents,

citicoline has the potential not only to stabilize the size or

locale of the area of damage, but also to contribute to the repair

of the damaged area.

Without being limited by theory, it is believed that citicoline has

at least a dual mechanism of action: limiting nerve damage and

further progression of disease and aiding in the repair of damaged

neuronal tissues. Administration of citicoline is believed to limit

the extent of the tissue damage by preventing the accumulation of

toxic free fatty acids. In addition, following its administration,

it is believed that citicoline is broken down into components,

including cytidine and choline, which are substrates required in the

formation of phosphatidylcholine, the primary phospholipid of nerve

cell membranes, via the Kennedy pathway. It is further postulated

that to normalize brain and/or muscle function, nerve cells damaged

by motor neuron or demyelinating diseases must manufacture new

membrane elements. As described below, in preclinical animal models

of ALS and MS, administration of citicoline is shown to reduce the

functional deficits produced by nerve degeneration.

Citicoline is preferably administered orally as a pharmaceutically

acceptable salt. The preferred salt is the monosodium salt of

citicoline, as this form is readily available in pharmaceutically

acceptable purity. Citicoline monosodium is an exogenous form of

cytidine-5'-diphosphocholine (CDP-choline). Endogenous CDP-choline

is a key intermediate in the biosynthesis of membrane

phosphatidylcholine, the primary lipid membrane component involved

in the dynamic regulation of cellular integrity.

Citicoline may be administered in the following daily dosages. All

dosages are provided on a citicoline monosodium basis and on a per

patient basis (ranging from about 45 kg to about 100 kg per patient

or 70 kg patient on average).

Generally daily citicoline dosages may range from about 100 mg to

about 5000 mg, desirably from about 250 to about 3000 mg and

preferably from about 500 to about 2000 mg. Doses may be

administered once or up to four or more times daily. A highly

preferred dosage is 500 mg administered twice per day per patient.

If greater therapeutic efficacy is required, a preferred

administration is 2000 mg administered in either a single 2000 mg

dose or two 1000 mg doses.

The treatment length is variable, but it has been observed that

patients tolerate citicoline well at doses ranging from about 250 mg

to about 2000 mg for prolonged periods, that is, from several weeks

to several years. Dosages may be varied over time depending on the

severity of symptoms, individual patient tolerance, route of

administration and response to treatment. Treatment may be continued

indefinitely if needed and if tolerated well.

Preferably, citicoline is administered orally in the form of

capsules, cachets, tablets or lozenges, or as a powder or granules

for reconstitution as a solution or suspension in an aqueous or non-

aqueous liquid. Administration may also be in the form of a bolus,

electuary, suppository, or paste. Formulations for inhalation, or

intranasal administration are also contemplated.

Formulations of the active ingredient, suitable for parenteral

administration, may comprise a sterile, aqueous preparation of the

citicoline active ingredient. The formulations may be presented in

unit dosage form and may be prepared by any of the methods well-

known in the art of pharmacology.

In addition to containing the standard and well known pharmaceutical

carriers and/or excipients, all of the above formulations may

contain other therapeutically active substances. Thus, the present

invention also contemplates a combination treatment regimen that

relates to the co-administration of citicoline and at least a second

therapeutic agent, or the respective pharmaceutically acceptable

salts thereof.

Broad categories of the at least second therapeutic agent are

contemplated. These agents include, but are not limited to,

glutamate and glycine antagonists such as neurontin, and drugs such

as ACTH, glucocorticoids (e. g., methylprednisolone, prednisone and

dexamethasone), antiinflammatory drugs, diphenylhydramine, quinine,

myotrophin, or IGF-1, BDNF, BFGF, beta-interferon, Betaseron,

Copaxone, Baclofen, Riluzole, and epitopes of myelin basic proteins

and the like, which are often used to treat ALS or MS.

Yet other therapeutic agents useful in combination with citicoline

are calcium channel blockers (e. g., AJ- 394, AK-275, Calpain

inhibitors, CD-349, Clentiaze, CNS-1237, CNS-2103, CPC-304 and CPC-

317, Dazodipine, Diperdinine, Emopamil, Fasudil, Lacidipine,

Lifarizine, Lomerizine, Magnesium, MDL : 28170, NB-818, Nilvadipine,

Nimodipine, NS-626 and related compounds, SM-6586, SNX- 111, S-312-

d, U-92032, UK-74505, US-035 and the like), agents targeted at

nitric oxide, agents targeted at various other neurotransmitters (e.

g., alpha2-receptor therapeutics, CV-5197, Dopamine receptors,

Enadoline, Lazabemide, Milnacipran, Nalmefene, RP-60180, SR- 57746A,

synaptic uptake blockers and the like), cytokines, hormones and

related products (e. g., AN- 100225 and AN-100226, Calcitonin gene-

related peptides, CEP-075 and related compounds, Ciliary

neurotrophic factor, Endothelial cell factor, Endothelin inhibitors,

FR-139317 Interleukin-1 receptor antagonist (lipocortin), JTP-2942,

Macrophage-regulating compounds, Motoneuronetrophic factor NBI-117,

Nerve growth factor, Neural stem cells, Neutrophil inhibitory

factor, NS-506, NT-3, Posatirelin, Schwann cell promoters, sCR1,

Somatomedin-1 and the like), free radical scavengers (e. g., EPC-K1,

MCI-186, Nicaraven, Phenazoviridin, Resorstatin, Rumbrin, Superoxide

dismutase, Tirilazad mesylate, U-88999E, Yissum project P-0619, YM-

737 and the like), gangliosides and related products (e. g., LIGA4,

LIGA4, Monosialoganglioside (GM1), ND-37, Siagoside and the like).

Still other classes of second therapeutic agents include, but are

not limited to, modulators of various specific enzymes,

neuroprotectives with " diverse " actions (e. g., Ademetionine sulphate

tosilate, Ancrod, Apocuanzine, CPC-111, CPC-211, HSV vectors, KF-

17329 and KF-19863, LY-178002, MS-153, Nicorandil, N-3393 and N-

3398, SUN 4757, TJ-8007, VA-045 and the like, and imaging or

contrast agents).

Therefore, a method is provided of treating a subject who is

suffering from motor neuron or demyelinating disease comprising co-

administering an effective amount of citicoline and at least a

second therapeutic agent, or their respective pharmaceutically

acceptable salts. The first dose may then be followed by the co-

administration of one or more subsequent doses of effective amounts

of citicoline alone, the at least second therapeutic agent alone, or

their respective pharmaceutically acceptable salts, or as subsequent

combinations thereof. Consistent with the other methods disclosed

herein, the first dose may be co-administered after diagnosis. By

the use of the term " co-administration, " is meant that the citicoline

and the at least second therapeutic agent, or their respective

pharmaceutically acceptable salts, are administered together or

sequentially.

The method using the contemplated combination therapy includes the

administration or co- administration of subsequent doses, which is

preferably carried out over a period of at least about 30 days.

In a specific embodiment of the invention, the co- administration of

subsequent doses is carried out over a period of at least about 4-8

weeks, preferably over a period of at least about six months to

about one year.

Furthermore, the first dose or subsequent doses are co-administered

one or more times daily over the predetermined period. It is

anticipated that subjects who may benefit the most from the

combination therapy are those who suffer from advanced ALS or other

motor neuron disease, or who are in the acute, active stage of a

demyelinating disease such as MS, or chronic progressive MS.

Maintenance doses may be required for some patients for the rest of

their lives.

In the composition, the effective amount of active ingredients in a

therapeutic dose may vary according to the particular need. Typical

ranges, however, may be from about 100 mg to about 5000 mg of

citicoline and about 10 mg to about 1000 mg of at least a second

therapeutic agent.

The present invention is illustrated by the Examples that follow, it

being understood, however, that the invention is not limited to the

specific details of these Examples.

Example 1 Experimental Allergic Encephalomyelitis (EAE) Model for MS

Experimental allergic encephalomyelitis (EAE) is an inflammatory

autoimmune demyelinating disease which can be induced in laboratory

animals by injection of myelin basic protein (MBP) or ground spinal

cord from another species. This artificially induced disease has

become the standard laboratory model for studying clinical and

experimental autoimmune diseases. There are many similarities

between EAE in animals and MS in humans, including chronic relapse.

Thus, EAE is a good predictor of efficacy of drugs and drug

combinations for treatment of various autoimmune diseases. Also,

because of the similarity in motor symptoms, EAE may also be

predictive of drug efficacy for ALS.

The EAE test model is employed to establish the activity of

citicoline against MS. Such testing is conducted according to the

following procedure.

Thirty female rats are injected in their foot pads with guinea-

pig spinal cord homogenate in complete Freund's adjuvant. The rats

are divided into three groups of 10 each. One group is administered

citicoline i. p. daily in a dose of 500 mg/kg beginning at 9 days

after inoculation. A second group is administered dexamethasone in

daily doses of approximately 0.0375 mg/kg s. c., beginning 9 days

after inoculation. The third group is a control to which is

administered 0.9% saline solution beginning 9 days after

inoculation. Duration of the treatment is approximately 17 days.

Animals are examined daily, which consists of weighing and scoring

for symptoms of EAE according to a disability scale of 0-4.

The results of such rat studies are shown in Figure 1. They

establish that citicoline inhibits the progress of EAE, with a dose

of 500 mg/kg exhibiting desirable levels of activity. Dexamethasone

at a relatively high dosage also inhibits the progress of EAE

initially.

Example 2 This example demonstrates the benefits of prevention

therapy with citicoline. Thirty-two female rats are divided

into four groups of eight each and on day one administered saline,

citicoline (500 or 1000 mg/kg, i. p.), or dexamethasone (0.0375

mg/kg), respectively. Dexamethasone acts as a positive control

through its ability to suppress immune function. On day two,

experimental autoimmune encephalomyelitis is induced in all rats by

injecting ground guinea pig spinal cord in complete Freund's

adjuvant in the foot pad. Therapy is continued daily. The rats are

tested daily and assigned a functional score of 0 to 4, with 0 being

normal and 4 representing death or inability to move. The results,

which are summarized in the Table, indicate that administration of

citicoline markedly reduces the rapid deterioration of motor

function in the test subjects compared to the saline group.

Numerical values are averages of the functional scores in each

treatment group from Days 14-21 of the study.

Table Mean Punctional Score Group Day Day Day Day Day Day Day Day 14

15 16 17 18 19 20 21 vehicle 0.72 0.94 1.0 1.12 1.16 1.19 1.35 1.25

(saline) Dexamethasone 0.14 0.42 0.0 0.14 0.28 0.64 0.57 0.64

(0.0375 mg/kg) citicoline 0.43 0.41 0.37 0.38 0.28 0.59 0.68 0.87

(500 mg/kg) citicoline 0.56 0.78 0.84 0.69 0.62 0.91 0.75 0.84 (1000

mg/kg) Example 3 This example demonstrates the benefits of

prevention therapy with a combination of citicoline and

dexamethasone. Sixteen female rats are divided into eight

groups of two each and on day one each group is administered one of

the following eight solutions: 1. Dexamethasone (Dex) 9.375 Fg/kg 2.

Dexamethasone 4.6 Fg/kg 3. Dexamethasone 2.3 Fg/kg 4. Citicoline 500

mg/kg + Dex 9.375 Rg/kg 5. Citicoline 500 mg/kg + Dex 4.6 Fg/kg 6.

Citicoline 500 mg/kg + Dex 2.3 Fg/kg 7. Citicoline 500 mg/kg 8.

Saline On the same day, experimental autoimmune encephalomyelitis is

induced in all rats by injecting ground guinea pig spinal cord in

complete Freund's adjuvant in the foot pad. Therapy is continued

daily. The rats are tested daily and assigned a functional score of

0 to 4, with 0 being normal and 4 representing death or inability to

move. The results, which are summarized in Figure 2, indicate that

administration of citicoline in combination with 2.3 Fg/kg of

dexamethasone reduces deterioration of motor function in the test

subjects to a level equal to or better than that observed for

administration of dexamethasone alone at levels up to 9.375 Rg/kg.

This enhancement of the activity of dexamethasone allows the same

results to be obtained without higher doses of dexamethasone, which

have the potential for toxic effects, as described in Goodman &

Gilman's The Pharmacological Basis of Therapeutics, Ninth Edition

(1996) at pages 1474-1476.

In addition, administration of citicoline in combination with 9.375

Fg/kg of dexamethasone reduces deterioration of motor function

substantially compared to the other treatments illustrated in Figure

2. This increased activity is achieved without exposing the subject

to the detrimental effects of higher steroid dosages.

Example 4 This example demonstrates the benefits of post- symptom

therapy with a combination of citicoline and dexamethasone. Eight

female rats are divided into four groups of two each and on

day one experimental autoimmune encephalomyelitis is induced in all

rats by injecting ground guinea pig spinal cord in complete Freund's

adjuvant in the foot pad. At symptom onset at approximately day 10,

each group is administered one of the following four solutions: 1.

Dexamethasone (Dex) 37.5 Rg/kg 2. Citicoline 500 mg/kg + Dex 9.375

Rg/kg 3. Citicoline 500 mg/kg 4. Saline, 0.9% (1 ml/kg, ip) Therapy

is continued daily. The rats are tested daily and assigned a

functional score of 0 to 4, as follows: 0: normal 0.1: tail weaker

than normal 0.25: tail fails to curl around examiner's finger 0.75:

tail strength only at base 1.0: loss of all tail strength 1.5: limp

tail + failure of one or more hind limbs to grip rotorod 1.5: hind

limb dragging (weak movement possible) 1.75: hind limb dragging +

failure of one or more hind limbs to grip rotorod 2.0: hind limb

paralysis 3.0: hind limb paralysis + fail rotorod test 4.0: total

paralysis or death The results, which are summarized in Figure 3,

indicate that administration of citicoline in combination with 9.375

Rg/kg of dexamethasone reduces deterioration of motor function in

the test subjects to a level equal to or better than that observed

for administration of dexamethasone alone at a relatively high level

of 37.5 Rg/kg. This enhancement of the activity of dexamethasone

allows the same results to be obtained without higher doses of

dexamethasone in a post-symptom treatment regimen. In addition,

administration of citicoline in combination with 9.375 Rg/kg of

dexamethasone reduces deterioration of motor function substantially

compared to administration of citicoline alone. This increased

activity is achieved without exposing the subject to the detrimental

effects of higher steroid dosages.

ExamRle 5 ALS Animal Model Twenty male three month old, transgenic

mice expressing familial ALS-linked mutations in the cytoplasmic

enzyme SOD1 are divided randomly into two groups of ten mice each:

ten animals treated with 500 mg/kg citicoline administered i. p.

five days a week and ten control animals treated with water.

The mice show progressive weakness arising from selective motor

neuron death, perikaryal proximal axonal swelling, axonal

degeneration, and severe skeletal muscle atrophy, all symptoms

consistent with familial ALS in humans.

After a trial period of about three months, improvements in the

motor behavior of treated animals and enhanced survival over control

animals is observed.

Generally, stabilization of the debilitating symptoms is achieved.

In some cases improvements are highly dramatic over the control

group.

Examples 6-13-Human Studies Example 6 A study of two sets of four

patients each with chronic multiple sclerosis is undertaken. Each

patient is first examined for normal hepatic, renal, and bone marrow

functioning to establish baseline values. Each of the patients in

each group is then treated either with citicoline dissolved in

sterile preservative-free isotonic saline 10% or oral tablet or

capsule. The citicoline is administered orally or intravenously at a

dosage of 250,500, or 1,000 mg each patient each day for six months.

Patients are examined on a daily basis. During the treatment period,

daily blood counts and twice weekly blood chemistries are performed

on each patient. The neurologic function of each of these patients

is measured using the expanded Krutzke disability status scale

(EDSS), and the Scripps neurologic rating scale (SNRS).

There is no evidence of any significant toxic side effects. None of

the eight patients exhibit any nausea, vomiting, skin rash, or

hepatic or renal dysfunction.

In essence, there is no evidence of toxicity in these eight patients

with normal marrow, hepatic and renal function. Likewise, the side

effects of the citicoline are imperceptible in these eight patients.

Measurement of neurologic function using the EDSS and SNRS scales

provides evidence of improvement in MS patients during treatment

with citicoline.

Example 7 Six patients with bulbar palsy caused by ALS are treated

with 500-1000 mg each citicoline by injection (monosodium salt

dissolved in 10 ml sterile isotonic solution of sodium chloride) or

by a solid tablet or soft gel capsule orally and administered daily.

As a result of treatment, remarkable improvements in bulbar symptoms

are observed, all six patients (three in each group) being able to

speak after 21 days of treatment. The side effects of this treatment

are insignificant and can be neglected in consideration of the

improvement obtained. Treatment is continued for 60 days and all six

patients exhibit improvement for several days after treatment is

halted.

Example 8 A double-blind study against placebo is performed as

follows: 77 patients suffering from ALS are treated with 500 mg each

citicoline by injection (monosodium salt dissolved in 10 ml sterile

isotonic solution of sodium chloride) or be a solid tablet or soft

gel capsule orally and administered daily, for a period of 12 to 18

months, and 78 patients receive a placebo.

The results obtained are analyzed in terms of survival in the study,

it being understood that " study drop-outs " (departures from the

study) include individuals who actually die, and also individuals

whose clinical state necessitates a tracheotomy or transfer to

assisted ventilation.

In this study, 50% of patients on placebo die, whereas this

percentage drops to 20% in patients on citicoline.

The probability in Wilcoxon's test (R. L.

PREUCTICE, Bioimetrika, 65,167-179 (1978)) is equal to 0.02 and the

probability in the stratified log-rank test (R. PETO and J. PETO,

Journal of the Royal Statistical Society, series A, vol. 135,185-207

(1972)) is equal to 0.09.

In subjects suffering from ALS with early bulbar involvement or the

bulbar form of the disease (the most serious form of the disease;

the usual mean survival of this type of patient is less than 3

years), 65% of patients on placebo die, whereas this percentage

drops to 30% in patients on citicoline. The probability in

Wilcoxon's test is equal to 0.020 and the probability in the log-

rank test is equal to 0.047.

Example 9 Six patients ranging in age from 33 to 38 who have

suffered from multiple sclerosis for more than 6 years and have been

bedridden for at least 4 months are treated with steroids after

which walking is possible with the aid of a walker. A wheelchair is

needed for travel outside the home. The attending physician advises

the patients that without chemotherapy (including treatment with

cytoxan), the patients would return to the bedfast state within 6

months.

The patients elect to discontinue all prescribed therapy and begin

taking approximately 1000 mg oral citicoline daily. The patients

observe no change in the status of the disease until about 10 weeks

have elapsed at which time they report feeling better than at any

time in the previous year. Within two to four additional weeks, all

patients are able to walk longer distances.

Example 10 Four patients with a history of leg weakness, progressive

loss of voice and multiple other neuromuscular symptoms are

diagnosed with ALS. The patients start on 250 mg/day oral citicoline

and an extensive exercise program.

Three months after therapy initiation, the condition of all six

patients deteriorates slightly, including progressive weakening of

the voice. However, the worsening of condition is minimal compared

to baseline ALS progression.

Example 11 Five patients ranging in age from 31 to 47 are diagnosed

with Charcot-Marie Tooth disease. The diagnosis is confirmed by

genetic studies revealing a duplicate locus on chromosome 17pll. 2

containing a gene for a peripheral myelin protein. All five patients

exhibit weakness and wasting of skeletal muscles as well as sensory

changes. Three are bedridden while the other two are confined to

wheelchairs.

The patients discontinue all prescribed therapy and begin taking 750

mg oral citicoline daily. No improvement is observed for 12 weeks,

after which progressive improvement is observed in each case.

After an additional four weeks of therapy, the bedridden patients

are able to use a wheelchair while the previously wheelchair bound

patients can use a walker for short distances.

Example 12 Seven patients ranging in age from 12 to 22 are diagnosed

with Friedreich's ataxia. All patients exhibit pes cavus

(foreshortening of the feet) with cocking of the toes and

unsteadiness in walking. The two oldest patients also exhibit

dysarthria.

The patients discontinue all prescribed therapy and begin taking 500

mg oral citicoline daily. After four weeks, progressive improvement

is observed in each case. All patients exhibit greater stability in

walking and the two oldest patients show improvement in speech.

Example 13 A. H. is a 54 year-old white, right-handed, married

female who presents with chronic progressive multiple sclerosis. The

patient reports first known neurological event to have occurred at

28 years old.

She described an optic neuritis event of her left eyes.

She was told at that time that she could have one of a variety of

medical conditions, including multiple sclerosis. She was given

prednisone by a general practitioner, and her optic neuritis

syndrome appeared to have resolved. She remained symptom-free for

approximately ten years.

At age 38, she described the recurrence of optic neuritis. She was

more descriptive of that event where the vision in the center of her

eye appeared distorted.

Peripheral vision was apparently unaffected.

Initially, she pursued the care of an ophthalmologist who felt the

etiology was once again related possibly to multiple sclerosis, She

went on to pursue further evaluation of an internist who later

referred her to a neurologist. At the time, she was described as

having relapsing-remitting multiple sclerosis. She received a course

of ACTH therapy. Initially, the symptoms had resolved, but she went

on to describe exacerbations of ophthalmological symptoms between

1982 and 1996, alternating between the right and left eyes. In 1986,

she described the onset of " falls. " She recalls being affected by the

dragging of her left foot on occasion and " stubbing her toe

frequently " on the ground. Her right lower extremity was notably

stronger than the left. She continued to have a gradual decline from

that point on and had no further symptom-free periods.

Since around 1986, she has received the following course of

treatments and therapies: Prednisone (worked initially),

Methylprednisolone (marginal effects), ACTH (worked initially),

Imuran (no effect), Cytoxan (uncertain of effect as course of

treatment was discontinued due to adverse events), Betaseron (no

effect), Cladribine (completed in January of 1998, and felt to have

had no effect), Non-allergenic diet (avoidance of dairy products and

other foods-no effect), Bee venom therapy (felt to have increased

the strength in legs, through this subsided after 3-4 months)

Currently, she is diagnosed with chronic- progressive multiple

sclerosis. She is afflicted with visual impairments, chronic

fatigue, difficulty with speech articulation, tremors in the right

and left hands, decreased fine motor movement in the right greater

than left hands and decreased motor strength in her lower

extremities. She is without weight bearing and is confined to a

wheelchair. When asked what her major barriers are as far as

functional abilities on a day to day basis, she places fatigue at

the top of her impairment list followed by speech difficulties and

decreased fine motor movement of her hands. She experienced marked

spasticity of her lower extremities with the progression of her

neurological condition over the course of several years and

eventually resorted to a baclofen pump which was implanted in 1997.

She has had continuous infusions into her spinal fluid since this

time, which has reduced the spasticity considerably in her lower

extremities. She receives 85 micrograms of baclofen per day. While

the baclofen has ameliorated the spasticity, it is felt that this

treatment has lessened her ability to bear weight on her lower

extremities.

Citicoline Response April 5,1998-The patient started on citicoline.

She began her treatment at 500 mg per day every morning for the

first two weeks. She reported no adverse side effects during this

time as well as no notable therapeutic effects. She and her husband

maintained a diary to record medication, potential adverse event and

neurological changes.

April 22,1998-Dosage was increased to 1000 mg per day (500 mg in the

a. m. and 500 mg in the p. m.). On the first two days of increased

dosage, the patient noted that her balance had been poor and that

her fatigue had increased in the early hours of the morning (not her

usual time for worsening of fatigue during the day). After two days,

however, the patient experienced increased endurance to stay up

later into the evenings (normal bedtime was previously 10 p. m.;

since the increased dose on citicoline, the patient found herself

retiring between midnight and 1: 00 a. m.). She was without those

early morning fatigue episodes noted on the first few days of

therapy, and, in general, was feeling relatively well.

Previously, she was discouraged from using the telephone in that her

speech articulation had been poor and perhaps her motivation use the

phone had lessened.

However, during this period of time, she also found herself both

willing and able to conduct telephone inquiries such as to conduct

banking activities. She described having a complicated discrepancy

with the bank and felt both motivated and capable of articulating

the situation to the bank successfully.

May 11,1998-Dosage was increased to 2000 mg per day (1000 mg in the

a. m., 1000 mg in p. m.) Once again, the patient noticed worsening

of fatigue in the morning hours for approximately two days, then

returned back to her baseline state of fatigue (generally less

fatigue in the morning and a gradual increase in fatigue as the day

processed). Once again, it was notable on citicoline (2000 mg/day)

that the patient was able to stay up for longer periods during the

day. She made a notation in a personal diary that on May 14th she

was making several phone calls and described an overall feeling of

wellness.

June 5,1998-The patient left for a 4 hour car drive to an out-of-

state family function. She recalled an inability to endure long car

rides for quite some time, and has avoided talking a car trip of

this duration in the past five years due to her subsequent

exhaustion following these events. She felt well upon arrival

despite the long drive. On June 6th, she forgot to take her

citicoline dose throughout the day. She described feeling " so-so and

not as well as the day before. " She also missed taking her citicoline

dose on the morning of June 7th. She detected a marked change in her

physical well-being in the sense that she was not as " up " and

attributed this to the return drive back to her home. They arrived

back at her house at approximately 2: 30 p. m. The patient described

noticing a pronounced difference in her overall state with mainly

increased fatigue. She described her tremors being worse, making

eating more difficult and she felt, in general, that she had to be

more careful as her motor and visual judgment had been off. She took

her evening dosage of citicoline (1000 mg) and on the following

morning she felt that she was back to baseline while on the drug.

At the end of June, A. H. took a cruise to Alaska with her family,

enduring the flight to and from the west coast. She tolerated the

trip and extensive traveling quite well.

Her neurologist has no knowledge that she is currently taking

citicoline. Prior to taking this medication, she was last seen by

her neurologist on April 21,1998. She was seen recently on July

7,1998.

The neurologist reported noticing an improvement in the patient's

eyes in that the " eye movement had improved. " The neurologist also

noticed that the strength with hand resistance had improved as well.

The patient continues to be on citicoline at a dose of 2000 mg per

day.