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CO-CURE Medical & Research Posts Only Digest - 11 Sep 2006 to 18 Sep 2006 (#2006-43)

There are 15 messages totalling 3527 lines in this issue.

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Date:    Tue, 12 Sep 2006 07:06:03 -0400
From:    "Bernice A. Melsky" <bernicemelsk@xxxxx.xxx>
Subject: RES: Changes in salivary cortisol and corticosteroid  receptor-alpha mRNA expression following a 3-week multidisciplinary  treatment program in patients with fibromyalgia

Changes in salivary cortisol and corticosteroid receptor-alpha mRNA
expression following a 3-week multidisciplinary treatment program in
patients with fibromyalgia.

Psychoneuroendocrinology. 2006 Sep 6; [Epub ahead of print]

Bonifazi M, Lisa Suman A, Cambiaggi C, Felici A, Grasso G, Lodi L,
Mencarelli M, Muscettola M, Carli G.

Dipartimento di Fisiologia, Via Aldo Moro, Universita degli Studi di Siena,
I-53100 Siena, Italy.

PMID: 16962248


The aim of the present study was to investigate the effects of a 3-week
residential multidisciplinary non-pharmacological treatment program
(including individually prescribed aerobic exercise and
cognitive-behavioral therapy) on fibromyalgia symptoms and
hypothalamic-pituitary-adrenal (HPA) axis function.

Salivary and venous blood samples were collected from 12 female patients
with fibromyalgia (age: 25-58) the day before and the day after the
treatment period: saliva, eight times (every two hours from 0800 to 2200h);
venous blood, at 0800h. Peripheral blood mononuclear cells (PBMC) were
separated and analyzed for glucocorticoid receptor-alpha (GR-alpha) mRNA
expression by semi-quantitative RT-PCR, while the salivary cortisol
concentration was determined by RIA. At the same time, pain and aerobic
capacity were evaluated.

Aerobic capacity improved at the end of the treatment program. The slope of
the regression of salivary cortisol values on sampling time was steeper in
all patients after treatment, indicating that the cortisol decline was more
rapid. Concomitantly, the area under the cortisol curve 'with respect to
increase' (AUC(i)) was higher and there was a significant increase in
GR-alpha mRNA expression in PBMC. The number of positive tender points,
present pain, pain area and CES-D score were significantly reduced after
the treatment, while the pressure pain threshold increased at most of the
tender points.

Our findings suggest that one of the active mechanisms underlying the
effects of our treatment is an improvement of HPA axis function, consisting
in increased resiliency and sensitivity of the stress system probably
related to stimulation of GR-alpha synthesis by the components of the treatment.

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Date:    Wed, 13 Sep 2006 11:20:01 -0400
From:    "Bernice A. Melsky" <bernicemelsky@xxxxx.xxx>
Subject: MED: Studies list more problems with pain-killing drugs

Studies list more problems with pain-killing drugs
Tue Sep 12, 5:32 PM ET - Reuters

Two studies offer more evidence about the dangers of some painkillers,
adding kidney problems to heart concerns already raised with the drug once
sold as Vioxx, researchers said on Tuesday.
One report from Boston's Brigham and Women's Hospital and Harvard Medical
School said an analysis of 114 studies involving more than 116,000 people
showed that rofecoxib (the chemical name for Vioxx) "was associated with
increased renal and (heart) arrhythmia risks."

Why the drug would cause kidney damage is unclear, it added.

Merck & Co Inc. withdrew Vioxx from the market in September 2004 after a
three-year study showed it doubled the risk of heart attack and strokes in
patients taking it for at least 18 months.

A second report from the University of Newcastle, New South Wales,
Australia, said a look at 23 studies confirms findings of an increased risk
of heart problems with Vioxx that could be found "during the first 30 days
of treatment. This conclusion is consistent with a recent reanalysis ...
which contradicts the original suggestion that the vascular risk was only
seen after 18 months."

The studies were published in this week's Journal of the American Medical
Association (JAMA) along with an editorial from David Graham, a physician
who works for the U.S. Food and Drug Administration but whose comments were
labeled as his own views and not those of the regulatory agency.

Merck is facing more than 11,500 product liability lawsuits from people
claiming to have been harmed by Vioxx.
"What the studies are going to do is provide substantial support for the
opinions being expressed by the plaintiffs' experts, and that will serve to
increase their credibility and persuasiveness to juries," said Frank
McClellan, professor at Temple University's Beasley School of Law.

"Those studies will also be used to cross-examine and impeach Merck's
experts who testify that there is no link between the drug and the
injuries," McClellan said. "The impact could be profound in the outcomes of
the trials."
Merck said it still believes the data confirm the increased heart risk
begins only after the medicine had been taken for 18 months.

Merck said in a statement the observations in the JAMA articles and
opinions expressed in an accompanying editorial regarding potential
increased risks with short-term Vioxx use "are not supported by the current
weight of clinical data."

The Australian analysis also found that celecoxib -- sold as Celebrex by
Pfizer Inc. -- was not associated with heart problems at a dose no greater
than 200 milligrams a day.

It said that its review "raises serious questions about the safety of
diclofenac, an older (analgesic) drug" which is sold more in Europe than
the United States.

"In conclusion ... diclofenac seems to share this risk and, unlike
celecoxib, it appears to be harmful at commonly used doses. We believe
there are grounds for reviewing its regulatory status," the report added.
In his editorial, Graham said the studies demonstrate that Vioxx "increases
the risk of acute myocardial infarction at low and high doses" and that
"there is no initial 18-month period of immunity from risk."
He said Celebrex increases heart risk at doses higher than 200 milligrams
per day and several other non-steroidal anti-inflammatory drugs (NSAIDs)
increase risk, including diclofenac, meloxicam, indomethacin and "probably"
ibuprofen, while studies agree naproxen is "neutral" for heart attack risk.
Graham added that for most patients with arthritis or other conditions
requiring chronic pain relief "naproxen appears to be the safest NSAID
choice from a cardiovascular perspective." Naproxen is commonly sold as
Aleve by Bayer Corp.

(Additional reporting by Bill Berkrot in New York)

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Date:    Wed, 13 Sep 2006 12:08:43 -0400
From:    "Bernice A. Melsky" <bernicemelsky@xxxxx.xxx>
Subject: RES: Comorbidity of Fibromyalgia and Psychiatric Disorders

Comorbidity of Fibromyalgia and Psychiatric Disorders.

J Clin Psychiatry. 2006 Aug;67(8):1219-1225.

Arnold LM, Hudson JI, Keck PE, Auchenbach MB, Javaras KN, Hess EV.

 From the Women's Health Research Program, the Department of Psychiatry,
University of Cincinnati College of Medicine, Cincinnati, Ohio (Dr. Arnold
and Ms. Auchenbach); Biological Psychiatry Laboratory, McLean Hospital,
Belmont, and the Department of Psychiatry, Harvard Medical School, Boston,
Mass. (Drs. Hudson and Javaras); Psychopharmacology Research Program, the
Department of Psychiatry, University of Cincinnati College of Medicine and
Mental Health Care Line and General Clinical Research Center, Cincinnati
Veterans Affairs Medical Center, Cincinnati, Ohio (Dr. Keck); the
Department of Biostatistics, Harvard School of Public Health, Boston, Mass.
(Dr. Javaras); and the Division of Immunology, the Department of Internal
Medicine, the University of Cincinnati College of Medicine, Cincinnati,
Ohio (Dr. Hess).

PMID: 16965199


OBJECTIVE: To assess the co-occurrence of fibromyalgia with psychiatric
disorders in participants of a fibromyalgia family study.

METHOD: Patients (probands) with fibromyalgia, control probands with
rheumatoid arthritis, and first-degree relatives of both groups completed a
structured clinical interview and tender point examination. The
co-occurrence odds ratio (OR) (the odds of a lifetime comorbid DSM-IV
disorder in an individual with fibromyalgia divided by the odds of a
lifetime comorbid disorder in an individual without fibromyalgia, adjusted
for age and sex) was calculated; observations were weighted by the inverse
probability of selection, based on the fibromyalgia status of the pro-band;
and standard errors were adjusted for the correlation of observations
within families. The study was conducted from September 1999 to April 2002.

RESULTS: We evaluated 78 fibromyalgia pro-bands and 146 of their relatives,
and 40 rheumatoid arthritis probands and 72 of their relatives. Among the
relatives of both proband groups, we identified 30 cases of fibromyalgia,
bringing the total number of individuals with fibromyalgia to 108, compared
with 228 without fibromyalgia. The co-occurrence ORs for specific disorders
in individuals with versus those without fibromyalgia were as follows:
bipolar disorder: 153 (95% CI = 26 to 902, p < .001); major depressive
disorder: 2.7 (95% CI = 1.2 to 6.0, p = .013); any anxiety disorder: 6.7
(95% CI = 2.3 to 20, p < .001); any eating disorder: 2.4 (95% CI = 0.36 to
17, p = .36); and any substance use disorder: 3.3 (95% CI = 1.1 to 10, p =
.040).

CONCLUSIONS: There is substantial lifetime psychiatric comorbidity in
individuals with fibromyalgia. These results have important clinical and
theoretical implications, including the possibility that fibromyalgia might
share underlying pathophysiologic links with some psychiatric disorders.

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Date:    Wed, 13 Sep 2006 16:59:54 +0200
From:    "Dr. Marc-Alexander Fluks" <fluks@xxx.xx>
Subject: RES,NOT: World famous researcher Professor declaring war on unhealthy fats

Quack science can be fun... below, please find today's commercial...


Source: Kilkenny Advertiser
Date:   September 13, 2006
URL:    http://www.kilkennyadvertiser.ie/index.php?aid=2685


Declaring war on unhealthy fats
-------------------------------

World famous researcher Professor Vasant Puri wants to see unhealthy trans fats
outlawed. Mention fats and most people automatically think of unhealthy ones
which we should try to eliminate or reduce in our diet. However, good fats also
exist and these play an important role in the functioning of our bodies.

Professor Basant Puri - a world renowned researcher and consultant at
Hammersmith Hospital in London who pioneered the use of natural fatty acids to
treat patients with chronic fatigue syndrome and depression and who visited
Ireland recently - believes trans fats or harmful fats in foods should be
outlawed in the same way as smoking in the workplace has been in this country.

Trans fats, also known as trans fatty acids, are a specific type of fat formed
when liquid oils are made into solid fats like shortening and hard margarine.
(A small amount of trans fats are found naturally, mainly in some animal-based
foods.)

Trans fats, which are considered to be the most harmful to people's health,
behave like saturated fat by raising low-density lipoprotein (LDL or 'bad')
cholesterol which increases your risk of coronary heart disease. They can be
found in some of the same foods as saturated fat, such as vegetable shortenings,
some margarines, crackers, sweets, biscuits, snack foods, fried foods, baked
goods, and other processed foods made with or fried in partially hydrogenated
vegetable oils.

'Ireland has been to the forefront in leading the way on many health issues,"
says Professor Puri. 'I believe that its decision to ban smoking in the
workplace has been an astounding success. I can personally verify this during my
recent visits to Dublin. As I'm a non-smoker, I visited a fine dining restaurant
and I was delighted to experience first-hand, the benefits of a smoke-free
environment. I also can recall their lead in banning hormones, thus removing
hormone-treated animals from the food chain. This gave them the edge in
marketing their beef across Europe free of residue.'

Now, he feels this country should take action against trans fats. 'I believe
Ireland should move with Denmark regarding these fats, if not banning products
with them at the very least introducing legislation requiring food manufacturers
to state the amount of trans fats in their products so that the public can make
an informed choice. I believe we should press for similar legislation in the
UK.'

Professor Puri - the author of over 25 books, including Chronic Fatigue
Syndrome: A natural way to treat ME and Natural Energy - Attention-Deficit
Hyperactivity Disorder (both published by Hammersmith Press) - advises people to
avoid trans fats if possible. He says they actively reduce the nutrient value of
essential fatty acids.

'With the high level of processed food now being consumed in the western
world, it is perhaps no surprise that diseases such as depression, chronic
fatigue syndrome and ADHD which are now so strongly linked with low levels of
essential fatty acids are on the increase.'

He says our bodies do not know how to deal with trans fats, some of which can
end up taking the place of 'good fatty acids' in cell membranes. This makes
cell membranes harder and more inflexible and may exacerbate brain diseases such
as attention deficit hyperactive disorder (ADHD), depression and ME. Trans-fats
are also likely to increase heart disease.

'The manufactured artificial trans-fats in our modern diets are probably doing
us a great deal of harm. They are found in many foodstuffs containing
hydrogenated vegetable oils, such as most cakes, pastries and biscuits and in
deep-fried foods and many margarines.'

How are the trans fats manufactured? 'To give one example when making
margarine, vegetable oils are industrially changed to turn them into solids at
room temperature. This process is known as hydrogenation and the result is
hydrogenated vegetable fat. Unfortunately the industrial processes involved
cause the fatty acids in the vegetable oil to be turned into strange molecules
called trans fats.'

Professor Puri recommends that people avoid products that have trans fats added
to them by food manufacturers. 'This does not mean you have to abstain from
all cakes, biscuits etc - alternatives include cakes and biscuits made with
butter rather than hydrogenated vegetable oil.'

He says people can help reverse some of the adverse effects of trans-fats by
taking a good omega supplement. He recommends one with a particularly high level
of EPA - eicosapentaenoic acid - an omega-3 fatty acid found in fish oil which
includes a special form of evening primrose oil, known as virgin evening
primrose oil. EPA has anti-viral properties and helps with sleep, also.

'The one supplement currently available to the general public that fulfils
these criteria is VegEPA. This is available via the internet by accessing the
website http://www.vegepa.com '

He says omega-3 and omega-6 fatty acids are very important to the proper
functioning of our bodies as they maintain the correct structure of cell
membranes.

The professor is calling on governments to take action in relation to trans
fats.

'Governments should act and introduce legislation requiring all food
manufacturers to state the amount of trans fats used in their products.'

What are trans fats?

Trans fats (also known as trans fatty acids) are a specific type of fat formed
when liquid oils are made into solid fats like shortening and hard margarine.

Where are they found?

Trans fat can be found in some of the same foods as saturated fat, such as
vegetable shortenings, some margarines, crackers, sweets, cookies, snack foods,
fried foods, baked goods, and other processed foods made with partially
hydrogenated vegetable oils.Trans fat behaves like saturated fat by raising
low-density lipoprotein (LDL or 'bad') cholesterol that increases your risk
of coronary heart disease (CHD).

How are they formed?

Trans fat are formed when hydrogen is added to vegetable oil, a process called
hydrogenation. This increases the shelf life and flavour of foods containing
these fats.

Are all fats the same?

No. While unsaturated fats (monounsaturated and polyunsaturated) are beneficial
when consumed in moderation, saturated fat and trans fats are not. Saturated fat
and trans fats raise LDL ('bad') cholesterol. Therefore, it is advisable to
choose foods low in both saturated and trans fats as part of a healthy diet.

--------
(c) 2006 Kilkenny Advertiser

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Date:    Wed, 13 Sep 2006 16:27:47 +0200
From:    "Dr. Marc-Alexander Fluks" <fluks@xxx.xx>
Subject: RES,NOT: Experts deny GWS

Source: Washington Post
Date:   September 13, 2006
Author: David Brown
URL:    http://www.thenewstribune.com/news/local/story/6092334p-5340677c.html


Experts deny 'Gulf War syndrome'
--------------------------------

WASHINGTON - An expert panel reviewing hundreds of studies has concluded there
is no single 'Gulf War syndrome' afflicting thousands of veterans of the
first war in Iraq, although they have suffered vague symptoms at a much higher
rate than other veterans.

They have also experienced post-traumatic stress disorder and depression two to
three times more frequently than other veterans, the panel found. Less certain
is a possibly higher risk for the neurological ailment Lou Gehrig's disease,
and possibly a rare birth defect in their children.

'We can't identify a Gulf War syndrome,' said Lynn Goldman, a physician and
epidemiologist who headed the 13-member committee appointed by the National
Academies Institute of Medicine.

Previous blue-ribbon panels, including several convened by the institute, one by
the Defense Department and one by the White House under Bill Clinton, reached
the same conclusion.

Tuesday's report is the latest in the important series, which the VA will rely
on to determine whether Gulf War veterans are eligible for special disability
benefits if they are found to suffer from illnesses that can be linked to their
service.

Veterans can now claim those benefits only by making an undiagnosed illness
claim, said Steve Robinson, a Gulf War Army veteran and government relations
director for Veterans for America.

'They keep saying it over and over, every year. We know that - we know that
there is no single thing that made veterans sick. We know this thing is likely a
combination of various exposures,' Robinson said in pushing for new studies he
hopes will find what ails tens of thousands of his fellow vets.

However, the report's confirmation that Gulf War veterans are sicker might
actually help them secure government benefits, said Shannon Middleton, assistant
director of health policy for the American Legion.

In preparing the report, Goldman and her colleagues read 850 studies done in
the years since soldiers in an Army Reserve unit in Indiana began reporting a
constellation of symptoms that included fatigue, joint and muscle pains,
difficulty concentrating and memory problems in the months after returning from
the Gulf in 1991. The panel gave more credibility to large, well-designed and
controlled studies; it did no original research of its own.

The largest and most representative study found that 29 percent of Gulf War
veterans reported physical complaints arising from several body systems,
compared to 16 percent of servicemen and women who were not sent to the Gulf.
This was not just an American phenomenon; British, Canadian, Australian and
Danish troops deployed to the Gulf also reported more symptoms than their
non-deployed counterparts.

The panel concluded that 'unexplained illnesses are the most prevalent health
outcome of service in the Gulf War.'

However, the array and severity of symptoms varies, and many are typical of
other conditions (such as fibromyalgia, multiple chemical sensitivity, and
chronic fatigue syndrome) whose cause is unknown.

'The nature of the symptoms suffered by many Gulf War veterans does not point
to an obvious diagnosis, etiology (cause), or standard treatment,' the panel
wrote.

Three studies examined by the panel found a slightly increased rate of
amyotrophic lateral sclerosis, also known as Lou Gehrig's disease, in Gulf
veterans. Overall, 107 cases of the fatal disease were found in about 700,000
men and women deployed. Goldman and her colleagues said this trend should be
followed with continued surveillance for the disease.

Among the 850 papers examined were several with less certain findings.

One found a slight increase in brain cancer among the 100,000 soldiers
theoretically exposed to sarin gas after rockets containing the nerve agent were
unwittingly destroyed in a large ammunition dump in Khamisiyah, Iraq, after the
war. That disease remained extremely rare, however, with only a handful of
tumors even in the 'exposed' group.

That finding is questionable for two reasons, however.

There is no evidence that any soldiers suffered actual exposure to sarin, which
causes well-recognized symptoms at very low concentrations. In addition, brain
tumors generally grow 10 to 20 years before being found. This study was done
nine years after Khamisiyah, suggesting that many of the tumors in the soldiers
had probably formed before the detonation.

One study also found a slightly increased rate of birth defects, especially of
the urinary tract, among Gulf war veterans. Another reported five cases of a
rare, disfiguring defect called Goldenhar syndrome in babies of Gulf veterans
conceived soon after the war, compared to only two in nondeployed veterans. The
children with Goldenhar were profiled in a Life magazine article that helped
popularize the idea that soldiers had been exposed to persistent toxins during
the war.

Because the number of defects was so small, however, these differences could
have easily happened by chance alone.

A big problem with evaluating the source of illness in Gulf veterans is that
little information was gathered on their exposure to pesticides, smoke and
battlefield contaminants - and even to vaccines and drugs administered by the
military.

The Associated Press contributed to this report.

--------
(c) 2006 Washington Post

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Date:    Thu, 14 Sep 2006 12:05:39 -0400
From:    Dr Marc-Alexander Fluks <fluks@xxx.xx>
Subject: NOT,RES: CFS patent John Gow et al.

Source: European Patent Office
Date:   August 10, 2006
URL:    http://v3.espacenet.com/textdoc?DB=EPODOC&IDX=WO2006082390&F=0
Ref:    http://www.me-net.dds.nl/forward/genomics.html


Bibliographic data of WO2006082390
----------------------------------

Publication number: WO2006082390
Publication date:   2006-08-10
Inventor:           GOW JOHN (GB);
                    CHAUDHURI ABHIJIT (GB)
Applicant:          UNIV GLASGOW (GB);
                    GOW JOHN (GB);
                    CHAUDHURI ABHIJIT (GB)
Classification:     international: C12Q1/68; G01N33/53; C12Q1/68; G01N33/53;
                    european: Application number: WO2006GB00332 20060201
                         Priority number(s): GB20050002042 20050201
View INPADOC patent family
Cited documents:    XP002361324
                    XP002377235
                    XP002377236
                    XP005062766
                    XP002377237


Abstract of WO2006082390
------------------------

The invention relates to materials and methods for diagnosis and treatment of
chronic fatigue syndrome/myalgic encephalitis. A number of genes are identified
which are expressed at abnormal levels in patients affected by CFS/ME as
compared to normal healthy individuals. These genes include those encoding
defensin a1, haemoglobin, CXCR4, tubulin beta 1, serine/threonine kinase 17B,
HLA DRss4 and prostaglandin D2 synthase. The genes identified provide objective
disease markers that may be used in diagnostic tests to support the diagnosis of
CFS/ME or for monitoring the effectiveness of therapy. They also provide a
rational basis for classifying CFS/ME patients according to the biochemical
lesion underlying their symptoms and enable provision of appropriate targeted
therapies.


Description of WO2006082390
---------------------------

[0001] Materials and Methods for Diagnosis and Treatment of Chronic Fatigue

[0002] Syndrome

[0003] Field of the Invention The invention relates to chronic fatigue
syndrome/myalgic encephalitis, and in particular to materials and methods for
its diagnosis and treatment.

[0004] Background to the Invention Chronic fatigue syndrome (CFS), also known
as Myalgic

[0005] Encephalomyelitis (ME), is an acquired disorder with long term
disability<1>. The illness affects both children and adults and is characterised
by persistent or relapsing fatigue of sufficient severity to interfere with
normal function. In addition, patients experience impairment in short term
memory and concentration, muscle pain and prolonged post-exertional malaise<2>.
The estimated overall prevalence of CFS/ME in the community is approximately
0.2-0.4% in adults and 0.07% in adolescents and children<3>.

[0006] The pathogenesis of CFS/ME is unknown. Diagnosis of CFS/ME according to
the internationally accepted definition (modified CDC criteria) is essentially
based on exclusion of known medical and psychiatric diseases<2>. As yet, there
is no specific or sensitive diagnostic test that positively establishes and/or
supports the clinical diagnosis of CFS/ME.

[0007] Nevertheless, in 2002, following the publication of a UK Government
Working Party report, Professor Sir Liam Donaldson confirmed that CFS/ME is a
debilitating and distressing condition affecting many people which should be
classed alongside other diseases such as multiple sclerosis and motor neurone
disease".

[0008] A significant proportion of patients report antecedent history of
community acquired viral or bacterial infections <4>'<5>'<6>. in addition, it is
clear that patients with CFS/ME have a hypoactive hypothalamic-pituitary-adrenal
(HPA) axis with altered neuroendocrine regulation affecting neurotransmitters
such as monoamines (norepinephrine, serotonin and dopamine) and acetylcholine in
the central nervous system<4>. Recent studies have demonstrated that viruses can
affect neurotransmitter functions<7>'<8>.

[0009] At the cellular level, fatigue has been linked with alterations in the
cell membrane ion-channel traffic and ATPase system<9>. ATPases are also linked
with neurotransmitter release (e. g. dopamine) <10> and cellular energy
metabolism via creatine phosphatase. Increased

[0010] ATPase activity has been reported in muscle biopsies from patients with
CFS <11>' <12>. Previous work<11>' <13> has raised the possibility that patients
with CFS may have an ion channel dysfunction. This dysfunction might be induced
by changes in the ion channel function, neurotransmitters involved in "gating"
the channel or by a shift in the balance of the cellular "energy charge",
i.e. the ratio between ATP and ADP that is normally a function of the ATPase
activity.

[0011] One previous study<43> has identified a number of genes differentially
expressed between PBMCs from CFS/ME patients and healthy controls. A number of
the genes identified are implicated in various aspects of immunological
function, from which the authors concluded that some kind of immunological
dysfunction may be involved in pathogenesis of CFS/ME. The authors were unable
to relate the observed expression patterns to any functional model of disease
etiology or pathology, and did not suggest that any of the genes which they
identified could serve as useful biomarkers for CFS/ME. Their conclusions are
consistent with previous studies which have suggested immunological abnormality
in CFS/ME (Refs. 44 to 48) but the precise mechanism of immunological
dysfunction in CFS/ME has not been established.

[0012] Summary of the Invention

[0013] The present inventors have identified a number of genes which are
expressed at abnormal levels in patients affected by CFS/ME as compared to
normal healthy individuals. In contrast to the earlier studies described above,
the present inventors have been able to use the expression patterns of these
genes to establish functional models of various aspects of the pathology of
CFS/ME, which explain many of the symptoms observed in affected individuals.
These provide a rational basis for classifying CFS/ME patients according to the
biochemical lesion underlying their symptoms and enable appropriate targeted
therapies to be provided for the first time.

[0014] The genes identified in the present invention provide obj ective disease
markers that may be used in diagnostic tests to support the diagnosis of CFS/ME
or in other applications. For example, the tests may enable deselection of
inappropriately diagnosed patients who have an alternative diagnosis for their
fatigue symptoms. The tests may also be used to classify patients according to
the particular biochemical basis for their symptoms. In turn, this may enable a
clinician to identify therapies which would be appropriate and rule out those
which would not. The tests may also be applied as an outcome measure for
interventional trials in CFS/ME, not least because no specific treatment is
currently considered to be effective in the patient population defined by the
clinical criteria. Further, the tests may offer support to CFS/ME patients for
their claims for disability and insurance. Finally, depending on the
specificity and sensitivity of the test, one may apply it to screen patients
with symptoms of chronic fatigue to identify or exclude CFS/ME. Given the fact
that chronic fatigue as a symptom is ten times commoner in the population (2%)
than CFS/ME, an estimated size of the market for a screening test is at least
10 million in the UK alone.

[0015] Thus the present invention provides a method for investigating whether a
test subj ect is affected by chronic fatigue syndrome/myalgic encephalomyelitis
(CFS/ME), the method comprising providing a biological sample from the subj ect
and determining the level in the sample of a biomarker for CFS/ME, wherein the
biomarker is an expression product of a gene shown in Table 1. Genes in Table 1
have been found to be overexpressed in CFS/ME compared to unaffected individuals.

[0016] The method may comprise determining the level, in the sample, of a
plurality of biomarkers (for example, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20
or more biomarkers), wherein each of the biomarkers is an expression product
of a gene shown in Table 1.

[0017] The method may comprise the step of determining the level in the sample
of one or more further biomarkers, being expression products of one or more
genes from Table 6. Indeed, an expression product from any one of the genes of
Table 6 may be used as a biomarker for CFS/ME, either alone or in combination
with other genes of Table 6 or of Tables 1 to 5.

[0018] The method may be considered to provide an expression profile of the one
or more biomarkers for CFS/ME, for the test subj ect, at the time of sampling.
By "expression profile" is meant a set of data relating to the level of
expression of one or more of the relevant biomarkers in a test subject, in a
form which allows comparison with comparable expression profiles (e.g. from
affected and/or unaffected individuals), in order to assist in the
determination of whether or not the subject is affected by CFS/ME.

[0019] The method typically involves correlating the results obtained with a
probability that the subj ect is affected by CFS/ME. In order to assist with
this correlation, the method of the invention may comprise the step of comparing
the expression profile for the test subj ect with one or more "reference"
expression profiles, that is to say one or more expression profiles
characteristic of unaffected subj ects (i.e. subj ects not suffering from
CFS/ME), and/or one or more expression profiles characteristic of affected subj
ects (i.e. subj ects not suffering from CFS/ME). Thus the level of
expression of one or more of the said biomarkers in the test subj ect is
typically compared with that characteristic of affected and/or unaffected
individuals.

[0020] The reference expression profiles may be profiles previously derived from
healthy or affected individuals, or may be artificial profiles which display
expression levels of the relevant biomarkers which are characteristic of the
relevant group. For example, they may be computer-generated from a plurality of
profiles previously derived from appropriate individuals. The profile (s)
characteristic of affected subj ects may be divided into subgroups, e.g.
according to their symptoms, as described in more detail below.

[0021] Taken alone, the expression profile for the test subj ect may not provide
an absolute diagnosis of CFS/ME. Normally, a clinician will also take account
of the physical and/or psychological symptoms of the subj ect in order to reach
a diagnosis. However the expression profile generated by the methods of the
invention provides useful data to help the clinician confirm or rej ect a
preliminary diagnosis based on physical and psychological symptoms alone. A
finding that one or more of these genes is upregulated in a particular
individual may therefore provide support for a diagnosis of CFS/ME.

[0022] By "upregulated" or "overexpressed" is meant that the gene in question
shows at least a two fold increase in expression at the level of r[alpha]RNA
and/or protein as compared to the level observed in unaffected (preferably
healthy) individuals. If desired, the level of overexpression required to
regard a result as positive may be set higher than this, e.g. a three, four,
five, six, seven, eight, nine or ten fold increase as compared to unaffected
individuals, or higher if required.

[0023] The genes in Table 1 not only provide biomarkers for CFS/ME, they also
provide the first evidence of specific biochemical pathways which may be
dysregulated in CFS/ME. They can be clustered into a number of subgroups,
based on their involvement in the same or related biochemical pathways whose
dysregulation is likely to play a role in either the underlying cause or the
symptoms of the disease state. Previously it has not been possible to identify
such pathways, which is a primary reason why no generally-recognised therapies
exist for CFS/ME. The subgroups of genes identified by the present inventors
provide an explanation for many of the symptoms displayed by those affected by
CFS/ME, suggest rational therapies for the condition, and provide means for
monitoring the efficacy of any therapy administered.

[0024] Without wishing to be bound by any particular theory, the inventors
believe that CFS/ME is not a genetic disease caused by a single or multiple gene
defects. Rather, they believe CFS/ME to be an acquired condition where there is
a shift in the functional systems of a select number of genes regulating
specific biological functions (e.g. infection and immunity, cell membrane
function and cell cycle) Based on the expression patterns they have observed,
they have established a model of "hub" and "network" genes which emphasises the
interrelation of these key genes, with the hub genes being the control centre
and the network genes being largely the effector arm of this functional system.
This set of hub and network genes defines the functional shift in the biological
systems of patients who continue to have symptoms due to CFS/ME.

[0025] Thus the genes of Table 1 are subdivided into "hub" genes and

[0026] "network" genes. Essentially, in any particular pathway the hub gene(s)
can be considered to lie upstream of the network genes, in that dysregulation
of the hub gene is likely to lead (directly or indirectly) to dysregulation of
the downstream genes. In Table 1, the hub genes are shown in section A, and the
network genes in section B.

[0027] Certain genes which appear to be highly significant (e.g. based on the
difference in their expression between the disease state and the normal state)
have been allocated to the "hub" group even where no related set of downstream
"network" genes is specifically identified.

[0028] Exemplary groups of genes identified in Table 1 are detailed below.

[0029] Defensin [alpha]1, CXCR4 and lactotransferrin (LTF) are all involved in
the response to intracellular infection. The network genes associated with
these hub genes include defensin [alpha]4, integrin [alpha]2B, integrin [beta]3,
arginase 1, arginase 2, thrombospondin 1, membrane associated protein 17
(MAP 17), Charcot Leyden Crystal Protein (CLC) and chondroitin sulphate
proteoglycan 2 (versican). These genes are shown in Table 2.

[0030] Haemoglobin Y (foetal haemoglobin) is part of the oxidative stress
response pathway, and is only expressed under conditions of oxidative stress.
Network genes associated with the oxidative stress pathway include other
haemoglobin genes including haemoglobin alpha 1, prostaglandin-endoperoxide
synthase 1 and prostaglandin- endoperoxide synthase 2. These genes are shown in
Table 3. However it will be noted that Table 6 includes genes for haemoglobins
alpha, beta, gamma and delta. Any one of the haemoglobin genes of Table 6 may
be used as a biomarker for CFS/ME, and may be used as well as, or in place of
haemoglobin gamma or alpha if desired. For simplicity, though, reference will be
made primarily to haemoglobin gamma.

[0031] Serine/threonine kinase 17B (STK17B) is implicated in apoptosis, as are
the network genes caspase 1, dynamin 1-like, and phosphatidyl serine binding
protein. These genes are shown in Table 4.

[0032] The MHC class II gene HLA-DR[beta]4 is upregulated in CFS/ME implying a
shift from antigen presentation by MHC class I to MHC II presentation.
Associated network genes include HLA-DQ[beta]1 and the immunoglobulin heavy
chain [gamma]3 (IgG3). These genes are shown in Table 5. IgG1 may be used as a
bior[alpha]arker as well as, or in place of, IgG3.

[0033] Methods for investigating whether a test subj ect is affected by CFS/ME
may therefore comprise determining the level of at least one gene from each of
at least two of Tables 2, 3, 4 and 5, and preferably from each of three or
four of said Tables. In preferred embodiments, at least one gene from each
said Table is a hub gene.

[0034] Preferred genes of Table 1 are defensin [alpha]1, haemoglobin [gamma],
CXCR4, tubulin beta 1 and HLA DR[beta]4. The method may involve testing
expression of any one, two, three, four or all five of these genes, and
optionally further hub genes of Table 1A.

[0035] Additionally or alternatively, the method may involve testing expression
of one or more of TSP 1, caspase 1 and/or IgG3.

[0036] The method may also comprise the step of determining the level of an
expression product of prostaglandin D2 synthase in sample. In contrast to the
genes of Table 1, prostaglandin D2 synthase is found to be downregulated in
individuals suffering from CFS/ME as compared to normal controls. Thus a
finding that prostaglandin D2 synthase is downregulated in a particular
individual may provide support for a diagnosis of CFS/ME.

[0037] By "downregulated" or "underexpressed" is meant that prostaglandin D2
synthase shows at least two fold higher expression at the level of mRNA and/or
protein in unaffected (preferably healthy) individuals than is observed in the
test subj ect. If desired, the level of downregulation required to regard a
result as positive may require the level found in unaffected individuals to be
three, four, five, six, seven, eight, nine or ten fold higher than in affected
individuals. The method may further involve the step of determining the level,
in a biological sample from the subj ect, of the peptide QYNAD, as described in
more detail below. The level of the peptide in the sample may form part of the
expression profile established for the subj ect. An elevated level of the
peptide as compared to unaffected individuals may be indicative of CFS/ME. By
an "elevated" level is meant at least twice the level found in unaffected
healthy controls, and preferably at least 5 times or at least 10 time the level
found in unaffected healthy controls.

[0038] It will be appreciated that individual patients who each satisfy the
criteria for CFS/ME may nevertheless present a range of very different symptoms.
Some of these can be explained by the groups of genes identified herein.

[0039] For example, CFS/ME has previously been suggested to involve immune
dysfunction. Some CFS/ME sufferers are particularly prone to infection by
viruses (e.g. influenza) and other pathogens; indeed some suffer recurrent
infections. Others are affected by atopic/allergic symptoms. These symptoms
may be associated with the apparent shift from Type I to Type II antigen
presentation by the MHC, which could impair the body' s ability to deal
efficiently with infections and could also exacerbate allergy/atopy.

[0040] Some CFS/ME sufferers describe their body as feeling prematurely aged,
and can display restricted mobility characteristic of much older individuals.
This might be explained by excessive apoptosis, particularly in the central
nervous system. Therefore patients having these symptoms might be expected to
show increased expression of serine/threonine kinase 17B (STK17B) and its
associated network genes.

[0041] Oxidative stress is also a cause of apoptosis, particularly in the
nervous system. Thus patients with increased expression of haemoglobin Y and its
associated network genes may also show, or be at risk of, accelerated programmed
cell death (increased level of apoptosis), particularly neuronal apoptosis.
Features of oxidative stress and neuronal apoptosis may include ageing,
cognitive impairment and chronic pain.

[0042] The genes of Table 2 are implicated in cellular protection ("defence")
against viral or bacterial infections. Thus an overexpressed defensin gene in
CFS/ME would be consistent with immune activation and correlate with symptoms of
recurrent influenza-type symptoms, sore throat and lymph node enlargement
experienced by patients with CFS/ME.

[0043] In a further aspect, the invention therefore provides a method for
classifying a subj ect affected by CFS/ME, the method comprising providing a
biological sample from the subject and determining the level in the sample of a
biomarker for CFS/ME, wherein the biomarker is an expression product of a gene
shown in one or more of Tables 2 to 5.

[0044] Preferably the biomarker is a hub gene from one of Tables 2, 3, 4 or 5.
More preferably, the method comprises determining the expression level of hub
genes from each of two, three or all four of Tables 2, 3, 4 or 5. The method
may also comprise determining network genes from one, two, three or all four of
Tables 2, 3, 4 and 5.

[0045] The individual can then be assigned to a subgroup of CFS/ME, dependent on
which group or groups of genes are found to be upregulated. Thus, for example,
the subj ect may be classified as being affected by one or more of oxidative
stress, excessive apoptosis, and immunological dysregulation (MHC I to II
shift). The assignment step may involve comparing the expression profile
obtained from the subj ect with one or more expression profiles characteristic
of individuals previously assigned to one or more subgroups of CFS/ME. It will
be appreciated that the classification may be performed using the same
expression profile as that established for determining whether the subj ect is
affected by CFS/ME.

[0046] Individuals in which these particular pathways are dysregulated may be
suitable for treatment by the methods described below. Therefore in a further
aspect the invention provides a method of determining whether an individual
affected by CFS/ME is suitable for treatment using such a therapy. The method
comprises determining whether the individual is affected by one or more of
oxidative stress, excessive apoptosis, and immunological dysregulation as
described above, and optionally prescribing a suitable treatment depending on
the outcome. These treatments are described in detail below.

[0047] Expression of individual biomarkers for CFS/ME (genes from Tables 1
to 5) may differ slightly between independent samples, leading to slightly
different expression profiles for individual samples. However, these particular
genes may provide a characteristic pattern of expression (expression profile) in
an affected individual (i.e. one suffering from CFS/ME) that is recognisably
different from that in an unaffected individual (i.e. one not suffering from
CFS/ME).

[0048] By creating a number of expression profiles from a number of known
affected and unaffected samples, it is possible to create a library of profiles
for both sample types. The greater the number of expression profiles, the
easier it is to create a reliable characteristic expression profile standard
(i.e. including statistical variation) that can be used as a control in a
diagnostic assay. Thus, a standard profile may be one that is devised from a
plurality of individual expression profiles and devised within statistical
variation to represent either the affected or unaffected profile. The
determination of the expression profile may be computerised and may be carried
out within certain previously set parameters, to avoid false positives and false
negatives.

[0049] The computer may then be able to provide an expression profile standard
characteristic of an affected sample and a normal sample. The determined
expression profiles may then be used to classify test samples as affected or
unaffected as a way of diagnosis.

[0050] Thus, in a further aspect the invention provides a method of creating a
library of expression profiles for use in determining whether an individual is
affected by CFS/ME, the method comprising

[0051] (a) providing biological samples from a plurality of individuals affected
by CFS/ME, and determining the level in each sample of one or more biomarkers
for CFS/ME to create a plurality of expression profiles from affected
individuals;

[0052] (b) providing biological samples from a plurality of individuals not
affected by CFS/ME, and determining the level in the sample of said one or more
biomarkers for CFS/ME to create a plurality of expression profiles from
unaffected individuals;

[0053] wherein the biomarkers comprise expression products of one or more genes
shown in Table 1.

[0054] Typically, the method comprises the step of retrievably storing each of
the expression profiles on a computer data carrier, in order to create a
database of expression profiles for both affected and unaffected individuals.

[0055] The invention further provides an expression profile database, comprising
a plurality of expression profiles of biomarkers for CFS/ME from affected and
unaffected individuals, wherein the biomarkers comprise expression products of
one or more genes shown in Table 1.

[0056] It will be appreciated that the profiles may be classified according to
their expression levels of the various groups of genes shown in Tables 2 to 5 as
already described. The combinations of biomarkers analysed in establishing the
expression profiles may therefore be chosen as described above.

[0057] The expression profiles may comprise data relating to the level, in
biological samples from the subj ects, of the peptide QYNAD. If required, the
methods may therefore comprise the step of determining the level of this peptide
in biological samples from the subjects.

[0058] At present there are no generally acknowledged treatments for CFS/ME.
One problem which has faced researchers attempting to develop and evaluate
suitable therapies is the lack of objective testable criteria to determine their
efficacy. The present invention provides a useful tool for monitoring the
efficacy of an interventional treatment for CFS/ME, by studying the effect of
the therapy, over time, on an individual' s expression profile of the biomarkers
described in Tables 1 to 5. The methods described may therefore be used to
evaluate the effectiveness of a test treatment, e.g. by testing it on a
population of subjects affected by CFS/ME, or to investigate an individual' s
response to a particular therapy, to see whether or not they are responding
appropriately.

[0059] Thus in a further aspect the invention provides a method of determining
the efficacy of a treatment for CFS/ME comprising the steps of:

[0060] (a) providing a biological sample from a subj ect affected by CFS/ME who
has been subj ected to said treatment, (b) determining the level in said sample
of one or more biomarkers for CFS/ME to create an expression profile for said
subj ect, and

[0061] (c) comparing said expression profile with

[0062] i) a comparable expression profile obtained from said test subj ect
before initiation of said treatment, and/or

[0063] ii) a comparable expression profile obtained from said test subj ect at
an earlier stage of said treatment, and/or

[0064] iii) a comparable expression profile characteristic of a subject who is
unaffected by CFS/ME,

[0065] wherein the one or more biomarkers for CFS/ME are expression products of
one or more genes shown in Tables 1 to 5. Preferred combinations of biomarkers
have already been described in relation to earlier aspects of the invention.

[0066] In general, a treatment may be considered to be effective if the subj
ect' s expression profile after treatment shows that the expression level of one
or more of the biomarkers is reduced compared to its level before treatment, or
its level earlier in a course of the treatment. Preferably the subj ect' s
expression profile after treatment approaches a profile characteristic of an
individual unaffected by CFS/ME. That is to say, the expression level of each
of the biomarkers falls within the normal range found in unaffected individuals.

[0067] Some treatments may be targeted to particular pathways known to be
dysregulated in CFS/ME. In such cases, it may be desirable to follow the
effect of the treatment on the expression levels of genes in the pathway likely
to be modulated by that treatment. For example, a treatment which is intended
to reduce oxidative stress, apoptosis or immune dysfunction (characterised by
MHC shift) may have a particular effect on the expression levels of the genes in
Tables 3, 4 and 5 respectively. The biomarkers chosen to study the effects of
the treatment may be chosen accordingly. Preferred combinations of biomarkers
from these Tables have already been described.

[0068] The method may further comprise determining the effect of the treatment
on the level of the peptide QYNAD in a biological sample from the test subject.

[0069] In the diagnostic and analytical methods described herein, the expression
levels of the chosen biomarker (s) for CFS/ME are preferably determined using
peripheral blood mononuclear cells (PBMCs) from the test subj ect. Therefore
the biological sample used to establish the expression profile is preferably a
blood sample or cells isolated from a blood sample. If desired, the method may
comprise the steps of enriching PBMCs in the sample, or isolating PBMCs from the
sample.

[0070] It will be understood that the methods described are generally performed
on a biological sample which has been isolated from the test subject. The
method may, but need not, comprise the actual step of isolating the sample from
the test subj ect, e.g. by taking a blood sample.

[0071] Typically, the expression level of each of the biomarkers is determined
by contacting the sample with a binding agent capable of binding specifically to
an expression product of the genes encoding that biomarker (a gene of Table 1, 2,
3, 4, 5 or 6). Binding between the agent and an expression product present
in the sample is then determined. If it is desired to determine the expression
level of more than one gene, then the sample may be contacted with a plurality
of binding agents, either simultaneously or sequentially, each binding agent
being capable of binding specifically and individually to an expression product
of one of the biomarker genes. Binding agents capable of binding to nucleic
acid expression products (e.g. mRNA, pre-mRNA) are typically nucleic acid
primers or probes having a sequence of, or complementary to, a portion of the
nucleic acid expression product. This enables the binding agent to hybridise
under suitable conditions with the nucleic acid expression product itself, e.g.
in a Northern blot or in situ hybridisation assay, or to a cDNA copy of the
nucleic acid expression product, e.g. in a RT-PCR assay, Southern blot or
microarray assay.

[0072] Binding agents capable of binding to polypeptide expression products
include ligands and receptors for the polypeptide in question. Particularly
preferred examples of binding agents are antibodies specific for (e.g. raised
against) the desired polypeptide, or fragments thereof comprising an antigen
binding site. These may be used in a variety of immunological assay methods,
including Western blots and ELISA assays, as well as in microarray assays.

[0073] It may be desirable to isolate expression products or particular
fractions thereof (e.g. total RNA, mRNA, total protein, soluble proteins or
membrane proteins) from the sample before contacting them with the binding agent.
The extent to which this is necessary will vary depending on the chosen assay
method.

[0074] Simple immunological assays (such as ELISA assays) or PCR-based assays
using ten biomarkers or less (e.g. 2, 3, 4, 5, 6, 7, 8, 9 or 10
biomarkers) are particularly preferred, because they can readily be adapted to
analyse large numbers of samples in a relatively short space of time, at
relatively low cost. Such assays are also well- suited to automation. Preferred
markers and combinations of markers are as described elsewhere in this
specification.

[0075] The diagnostic and analytical methods described above may also comprise
determining the level, in a biological sample from the subject, of the peptide
QYNAD. It is believed that the peptide may be found in serum and cerebrospinal
fluid (CSF) in most or all of the population, but it is found at elevated levels
in subjects suffering from inflammatory and immunological disorders of the
nervous system (such as multiple sclerosis and Guillain-Barre syndrome). The
present inventors have now established that it is also found at significantly
elevated levels in subj ects suffering from CFS/ME.

[0076] The biological sample used for determining the peptide level may be a
blood sample (or serum derived therefrom). Thus, conveniently, the same blood
sample may be used for determining levels of the peptide and the other
biomarkers of CFS/ME. Alternatively, a different sample may be used, in
particular, a sample of cerebrospinal fluid (CSF) where available.

[0077] The level of the peptide may be determined as described by Brinkmeier et
al.<16>, e.g. by gel filtration chromatography. Alternatively conventional
immunoassays such as ELISAs or Western blots may be used, employing antibodies
raised against the peptide.

[0078] The assay for the peptide is generally performed on a sample isolated
from the test subj ect. The method of the invention need not comprise the step
of actually isolating the sample from the test subj ect.

[0079] In a further aspect, the invention provides a kit for use in a diagnostic
or analytical method as described herein, the kit comprising a plurality of
binding agents, each capable of binding specifically and individually to an
expression product of one of the genes of Table I1 or the peptide QYNAD. Thus
the kit includes binding agents specific for expression products of two or more
genes of Tables 1, or at least one of the genes of Table 1 and the peptide QYNAD.
The binding agents provided in the kit may be capable of binding specifically
and individually to expression products of 2, 3, 4, 5, 6, 7, 8, 9, 10, 15,
20 or more of the genes of Table 1 and optionally the peptide QYNAD.

[0080] The kit is suitable for use in the methods of the invention described in
this specification, and may comprise instructions for performing one or more
methods of the invention.

[0081] The binding agents may be immobilised on one or more solid supports.
Discrete supports may each carry only one type of binding agent. For example,
distinct populations of beads may each carry one type of binding agent.
Alternatively, a single support may carry more than one type of binding agent.
Indeed, one support (e.g. a microarray chip) may carry all of the different
types of binding agent provided with the kit.

[0082] In addition, the kit may comprise one or more binding agents capable of
binding specifically to an expression product of a control gene which is not
differentially expressed between individuals affected and unaffected by CFS/ME.
The level of expression from this control gene may be measured m order to assist
in quantification of the expression products of the genes of Table 1, and/or
for quality assurance of an assay performed using the kit. Preferably a control
gene is chosen which is constitutively expressed in the cells of the biological
sample (i.e. always expressed, at substantially the same level, under
substantially all conditions). Such genes are often referred to as
"housekeeping" genes. Examples include glyceraldehyde phosphate dehydrogenase
(GAPDH), [beta]-actin, and abl (ableson tyrosine kinase).

[0083] The kit may comprise yet further binding agents capable of binding to
expression products of other biomarker genes or control genes. However, m
preferred embodiments, the kit comprises binding agents for expression products
of less than 1000 different genes, e.g. less than 500 different genes, less
than 100, less than 50, less than 40, less than 30, less than 20, or less
than 10 different genes. As explained above, the groups of genes identified by
the present inventors also suggest specific therapies for CFS/ME.

[0084] Patients displaying signs of oxidative stress may be treated with an
anti-oxidant. Thus the invention provides a method of treating CFS/ME in an
individual suffering therefrom, comprising administering an effective amount of
an anti-oxidant.

[0085] Therapeutically effective amounts of a corticosteroid and/or minocyline
may also be administered to the subject in conjunction with the anti-oxidant.
As will be appreciated, the individual active agents may be administered
individually (in two or more separate compositions) or together (in the same
composition).

[0086] The invention further provides the use of an anti-oxidant in the
preparation of a medicament for the treatment of CFS/ME. The medicament may be
formulated for administration in conjunction with a corticosteroid and/or
minocycline. Alternatively, the medicament may comprise a corticosteroid and/or
minocycline.

[0087] Examples of suitable anti-oxidants include coenzyme Q10 and inhibitors of
cyclooxygenase (COX) enzymes, particularly COX II enzymes, such as celecoxib (
4- [5- (4-methyl[rho]henyl) -3- (trifluoromethyl) - 1H-pyrazol-1-yl]
benzenesulfonamide).

[0088] Patients showing evidence of abnormal apoptosis may be treated with
minocyline, which is an inhibitor of caspase 1, shown here to be upregulated in
individuals affected by CFS/ME. Minocyline has previously been suggested for
treatment of various neurological disorders including stroke, multiple sclerosis,
spinal cord injury, Parkinson' s disease, Huntington' s disease and
amylotrophic lateral sclerosis - see Wee Young et al., Lancet Neurology, 2004,
744-751 for review. Thus the invention provides a method of treating CFS/ME
in an individual suffering therefrom, comprising administering an effective
amount of minocycline.

[0089] Therapeutically effective amounts of a corticosteroid and/or an
anti-oxidant may also be administered to the subject in conjunction with the
minocycline. As will be appreciated, the individual active agents may be
administered individually (in two or more separate compositions) or together
(in the same composition).

[0090] The invention further provides the use of minocycline in the preparation
of a medicament for the treatment of CFS/ME. The medicament may be formulated
for administration in conjunction with a corticosteroid and/or an anti-oxidant.
Alternatively, the medicament may comprise a corticosteroid and/or an
anti-oxidant.

[0091] Oxidative stress often gives rise to apoptosis. Therefore it may be
advisable to treat patients showing signs of oxidative stress with an apoptosis
treatment such as minocycline as a precautionary measure if they do not already
show upregulation of genes involved in apoptosis.

[0092] Patients showing signs of immune dysfunction, and particularly the Type I
to II shift of MHC expression, may benefit from treatment with a corticosteroid.
This would address the hypocortisolism previously reported in some CFS/ME
patients.

[0093] Thus the invention provides a method of treating CFS/ME in an individual
suffering therefrom, comprising administering an effective amount of a
corticosteroid.

[0094] Therapeutically effective amounts of minocycline and/or an antioxidant
may also be administered to the subject in conjunction with the corticosteroid.
As will be appreciated, the individual active agents may be administered
individually (in two or more separate compositions) or together (in the same
composition).

[0095] The invention further provides the use of a corticosteroid in the
preparation of a medicament for the treatment of CFS/ME. The medicament may be
formulated for administration in conjunction with minocycline and/or an
anti-oxidant. Alternatively, the medicament may comprise minocycline and/or an
anti-oxidant.

[0096] An example of a suitable corticosteroid is hydrocortisone. Others
include dexamethasone and prednisone.

[0097] The invention further provides pharmaceutical compositions suitable for
the treatment of CFS/ME. Thus there is provided a pharmaceutical composition
comprising a therapeutically effective amount of minocycline in combination with
a therapeutically effective amount of a corticosteroid and/or an anti-oxidant
and a pharmaceutically acceptable carrier.

[0098] Also provided is a pharmaceutical composition comprising a
therapeutically effective amount of a corticosteroid in combination with a
therapeutically effective amount of minocycline and/or an anti-oxidant, and a
pharmaceutically acceptable carrier.

[0099] Also provided is a pharmaceutical composition comprising a
therapeutically effective amount of an anti-oxidant in combination with a
therapeutically effective amount of a corticosteroid and/or minocycline, and a
pharmaceutically acceptable carrier.

[0100] As described above, a particular treatment may be determined for any
individual based on their expression profile of CFS/ME biomarkers. For example,
an individual showing upregulation of oxidative stress pathway genes may be
particularly suitable for treatment with an anti-oxidant. However it may not be
convenient or necessary to establish a suitable expression profile before
beginning treatment. Therefore it may be desirable to administer one, two or
all three of the treatment types described above to an individual affected by
CFS/ME without recourse to expression profiling.

[0101] Description of the Drawings

[0102] Figure 1 shows a RT-PCR assay of v-ATPase mRNA in PBMCs from three
healthy control individuals (lanes 2 to 4) and three patients with CFS/ME
(lanes 5 to 7). Lane 1 contains molecular weight markers.

[0103] Figure 2 shows a RT-PCR assay of Defensin alpha 1 mRNA in PBMCs from
three healthy control individuals (lanes 2 to 4) and three patients with CFS/ME
(lanes 6 to 8). Lanes 1 and 9 contain molecular weight markers.

[0104] Figures 3 and 4 show Western blots of proteins extracted from PBMCs from
healthy individuals and individuals suffering from CFS/ME, stained with
antibodies against Defensin alpha 1 and Thrombospondin 1 respectively. In each
case, lanes 1 and 12 contain molecular weight markers, lanes 2 to 11 contain
samples from patients with CFS/ME, and lanes 13 to 22 contain samples from
healthy control individuals. Purified protein extract from 2x10<5> cells was
loaded per lane. Proteins were separated on 10% SDS-PAGE gels and blotted onto
a nylon membrane.

[0105] Figure 5 shows a SDS-PAGE gel, stained with Coomassie blue, of the same
samples used to prepare the Western blots shown in Figures 3 and 4. Lanes 11 and
22 contain molecular weight markers, lanes 1 to 10 contain samples from
patients with CFS/ME, and lanes 12 to 21 contain samples from healthy control
individuals. As in Figures 3 and 4, protein extract from 2x10<5> cells was
loaded per lane.

[0106] Detailed Description of the Invention

[0107] Chronic Fatigue Syndrome/Myalgic Encephalomyelitis

[0108] CFS is typically diagnosed using the modified CDC criteria described by
Fukuda et al.<2>. All other conditions or diseases which could explain a
patient' s symptoms are first excluded. Having done this, CFS/ME is diagnosed
if the patient has been affected by 6 months or longer of persistent relapsing
or persistent fatigue accompanied by four or more concurrent symptoms including
impaired memory severe enough to affect normal daily function, sore throat,
tender lymph nodes, muscular or joint pain, new headaches, unrefreshing sleep
and post-exertional malaise lasting for more than 24 hours. For the purposes of
this specification, individuals satisfying these criteria are considered to be
affected by CFS/ME.

[0109] Assay methods

[0110] The genes identified in Table 1 provide biomarkers which may be used in
diagnostic assays to support, confirm, or refute a diagnosis of CFS/ME. With
the knowledge of this set of genes, it is possible to devise many methods for
determining a suitable expression profile of one or more CFS/ME biomarkers in a
particular test sample.

[0111] Typically, the method involves contacting expression products from the
sample with a binding agent capable of binding to an expression product of a
gene identified in Table 1. The expression product may be a transcribed nucleic
acid sequence or an expressed polypeptide.

[0112] The transcribed nucleic acid sequence may be mRNA or pre-mRNA.
Alternatively, the expression product may also be cDNA produced from said mRNA.
The binding member may a nucleic acid having a sequence complementary to that of
the RNA or cDNA which is consequently capable of specifically binding to the
transcribed nucleic acid or cDNA under suitable hybridisation conditions, e.g.
by Northern blotting, in situ hybridisation, or Southern blotting.

[0113] Such protocols may use probes of at least about 20-80 bases in length.
The probes may be of 100, 200, 300, 400 or 500 bases in length or more.
Binding assays may be conducted using standard procedures, such as described in
Sambrook et al., Molecular Cloning A Laboratory Manual (New York: Cold Spring
Harbor Laboratory Press, 1989 or later editions).

[0114] RT-PCR procedures (including quantitative PCR procedures) may also be
used to analyse the presence or amount of mRNA or precursor mRNA in a given
sample. A suitable primer having at least 15 to 20 bases complementary to the
desired mRNA or precursor mRNA sequence will typically be used to prime cDNA
synthesis. Alternatively a poly-T primer (optionally comprising one or more
random nucleotides at the 3' end) may be used to prime cDNA synthesis from all
mRNA in the sample.

[0115] Subsequently, a segment of the cDNA is amplified in a PCR reaction using
a pair of nucleic acid primers, each typically having at least 15 to 20 bases
complementary to the desired RNA sequence. The skilled person will be able to
design suitable probes or primers based on the publicly available sequence data
for the genes in question (see Table 1 for suitable accession numbers).

[0116] Where the expression product is the expressed polypeptide, the binding
member is preferably an antibody raised against or otherwise specific for the
desired polypeptide, or any other molecule comprising the antigen binding site
from such an antibody.

[0117] The skilled person will realise that other binding agents may be used as
appropriate. Suitable agents may include naturally- occurring ligands and
receptors for the desired polypeptide, aptamers, etc. For example, aptamers
are nucleic acid molecules (typically DNA or RNA), selected from libraries on
the basis of their ability to bind other molecules. Aptamers have been
identified which can bind to other nucleic acids (by means other than
conventional Watson-Crick base pairing), proteins, small organic compounds,
and even entire organisms. The binding agent (e.g. a nucleic acid probe or
antibody) may be fixed to a solid support. The expression products may then be
passed over the solid support, thereby bringing them into contact with the
binding agent. Conveniently, the binding agents are immobilised at defined,
spatially separated locations, to make them easy to manipulate during the
assay. The solid support may be a glass surface, e.g. a microscope slide,
beads, fibre-optics or microarray chip. In the case of beads, each binding
agent may be fixed to an individual bead and they may then be contacted with the
expression products in solution.

[0118] The sample is generally contacted with the binding agent (s) under
appropriate conditions which allow the analyte in the sample to bind to the
binding agent(s). The fractional occupancy of the binding sites of the
binding agent(s) can then be determined.

[0119] Whatever the chosen assay system, there are numerous ways to detect
interaction between the binding agent and the expression product (analyte) to be
determined, either by directly or indirectly labelling "the analyte or binding
agent, or by using a developing agent to arrive at an indication of the presence
or amount of the analyte in the sample. A developing agent may be a secondary
binding agent, capable of binding to a complex between analyte and primary
binding agent. For example, if a primary antibody is used as a binding agent,
the developing agent may be a secondary antibody capable of binding either to
the primary antibody, or to a different epitope on the analyte to that
recognised by the primary antibody.

[0120] Typically, the analyte, binding agent or developing agent is directly or
indirectly labelled (e.g. with radioactive, fluorescent or enzyme labels,
such as horseradish peroxidase) so that they can be detected using techniques
well known in the art. Directly labelled agents have a label associated with or
coupled to the agent, Indirectly labelled agents may act on a further species
to produce a detectable result. Thus, radioactive labels can be detected using
a scintillation counter or other radiation counting device, fluorescent labels
using a laser, confocal microscope, etc., and enzyme labels by the action of
an enzyme label on a substrate, typically to produce a colour change. In
further embodiments, the developing agent or analyte is tagged to allow its
detection, e.g. linked to a nucleotide sequence which can be amplified in a
PCR reaction to detect the analyte. Other labels are known to those skilled in
the art are discussed below.

[0121] The developing agent (s) can be used in a competitive method in which
the developing agent competes with the analyte for occupied binding sites of the
binding agent, or non-competitive method, in which the labelled developing agent
binds analyte bound by the binding agent or to occupied binding sites. Both
methods provide an indication of the number of the binding sites occupied by the
analyte, and hence the concentration of the analyte in the sample, e.g. by
comparison with standards obtained using samples containing known concentrations
of the analyte.

[0122] In alternative embodiments, the analyte can be tagged before applying it
to the support comprising the binding agent.

[0123] There is an increasing tendency in the diagnostic field towards
miniaturisation of such assays, e.g. making use of binding agents (such as
antibodies or nucleic acid sequences) immobilised in small, discrete locations
(microspots) and/or as arrays on solid supports or on diagnostic chips. These
approaches can be particularly valuable as they can provide great sensitivity
(particularly through the use of fluorescent labelled reagents), require only
very small amounts of biological sample from individuals being tested and allow
a variety of separate assays can be carried out simultaneously. This latter
advantage can be useful as it provides an assay employing a plurality of
analytes to be carried out using a single sample. Examples of techniques
enabling this miniaturised technology are provided in WO84/01031, WO88/1058,
WO89/01157, WO93/8472, WO95/18376/ WO95/18377, WO95/24649 and EP 0 373 203 A.

[0124] Other methods which do not rely on labelling techniques may also be used
to detect interaction between binding agent and reporter molecule, including
physical methods such as surface plasmon resonance, agglutination, light
scattering or other means.

[0125] Expressed nucleic acid (mRNA, pre-mRNA) can be isolated from the cells
using standard molecular biological techniques. The expressed nucleic acid
sequences corresponding to the gene or genes of Table 1 can then be amplified
using nucleic acid primers specific for the expressed sequences in a PCR, e.g.
real time PCR, multiplex PCR, etc. The skilled person will be able to select
or design a suitable reaction type and protocol depending on, e.g. the number
and particular combination of genes to be analysed. If the isolated expressed
nucleic acid is mRNA, this can be converted into cDNA for the PCR reaction using
standard methods.

[0126] The primers may conveniently introduce a label into the amplified nucleic
acid so that it may be identified. Ideally, the label is able to indicate the
relative quantity or proportion of nucleic acid sequences present after the
amplification event, reflecting the relative quantity or proportion present in
the original test sample. For example, if the label is fluorescent or
radioactive, the intensity of the signal will indicate the relative
quantity/proportion or even the absolute quantity, of the expressed sequences.
The relative quantities or proportions of the expression products of each of the
genes of Table 1 may be used to establish a particular expression profile for
the test sample.

[0127] Other methods for detection of nucleic acid expression products may also
be used, such as in situ hybridisation, Northern blot, etc. Likewise, protein
expression products may be detected by any suitable technique. Immunological
techniques are particularly preferred, in which antibodies specific for the
particular polypeptide gene product (s), are used as binding agents, although
other binding agents such as receptors or ligands capable of binding to the
proteins of interest may be employed.

[0128] In some embodiments, protein expression products from the sample under
test are immobilised on a solid phase and contacted with a binding agent
specific for one or more of the proteins of Table 1 under appropriate conditions
which allow binding between the protein and the binding agent. The amount of
the binding agent found at the surface is then determined. For example, the
binding agent may be directly labelled. Alternatively, the immobilised antibody
may be contacted with a labelled developing agent capable of binding to the
primary antibody. Examples of this type of assay include Western blotting, and
certain ELISA (enzyme-linked immunosorbent assay) techniques.

[0129] In other embodiments, a binding agent is immobilised on a solid phase
and contacted with the sample under suitable conditions to allow binding to take
place. The fractional occupancy of the binding sites of the binding agent (s)
can then be determined either by directly or indirectly labelling the analyte or
by using a developing agent or agents to arrive at an indication of the presence
or amount of the analyte in the sample.

[0130] An example of this type of assay is an antibody sandwich assay (e.g. an
ELISA), which employs two antibodies each capable of binding to a different
site on the biomarker protein. The first is immobilised on a solid phase for use
as the binding agent. After contact with the analyte, the second antibody is
used to detect complexes between the first antibody and analyte. Whichever
method is chosen, it is important that the assay provides a read-out of the
level of expression of the biomarker genes which allows results from different
individuals to be compared reliably with one another. By way of example, the
level of a particular expression product may be determined as a proportion of
the total expression products found in the sample. Alternatively, the level of
a particular expression product may be determined in relation to the level of
expression of a control gene such as a housekeeping gene, or the like.
Alternatively, it may be convenient to determine the absolute amount of a
particular expression product, e.g. by comparison with known standards. The
skilled person will be capable of designing a suitable protocol for any given
assay method, and will also be aware of other suitable embodiments.

[0131] Antibodies

[0132] It has been shown that fragments of a whole antibody can perform the
function of binding antigens. The term "antibody" is therefore used herein to
encompass any molecule comprising the binding fragment of an antibody, and the
term binding agent and binding site should be construed accordingly. Examples
of binding fragments are (i) the Fab fragment consisting of VL, VH, CL and CHl
domains; (ii) the Fd fragment consisting of the VH and CHl domains; (iii) the
Fv fragment consisting of the VL and VH domains of a single antibody; (iv) the
dAb fragment (Ward, E.S. et al., Nature 341, 544-546 (1989)) which
consists of a VH domain; (v) isolated CDR regions; (vi) F (ab ') 2 fragments,
a bivalent fragment comprising two linked Fab fragments (vii) single chain Fv
molecules (scFv), wherein a VH domain and a VL domain are linked by a peptide
linker which allows the two domains to associate to form an antigen binding site
(Bird et al, Science, 242, 423-426, 1988; Huston et al, PNAS USA, 85, 5879-5883,
1988). In preferred embodiments the binding agent comprises a single antigen
binding site specific for the analyte, i.e. a monovalent antibody or antibody
fragment.

[0133] Pha rmaceutical compositions Pharmaceutical compositions as described in
this specification typically comprise, in addition to one or more suitable
active agents, a pharmaceutically acceptable excipient, carrier, buffer,
stabiliser or other materials well known to those skilled in the art. Such
materials should be non-toxic and should not interfere with the efficacy of the
active ingredient. The precise nature of the carrier or other material may
depend on the route of administration, e.g. oral, intravenous, cutaneous or
subcutaneous, nasal, intramuscular, intraperitoneal routes.

[0134] Pharmaceutical compositions for oral administration may be in tablet,
capsule, powder or liquid form. A tablet may include a solid carrier such as
gelatin or an adjuvant. Liquid pharmaceutical compositions generally include a
liquid carrier such as water, petroleum, animal or vegetable oils, mineral oil
or synthetic oil.

[0135] Physiological saline solution, dextrose or other saccharide solution or
glycols such as ethylene glycol, propylene glycol or polyethylene glycol may be
included.

[0136] For intravenous, cutaneous or subcutaneous injection, or inj ection at
the site of affliction, the active ingredient will be in the form of a
parenterally acceptable aqueous solution which is pyrogen-free and has suitable
pH, isotonicity and stability. Those of relevant skill in the art are well able
to prepare suitable solutions using, for example, isotonic vehicles such as
Sodium Chloride Inj ection, Ringer ' s Inj ection, Lactated Ringer ' s Inj
ection. Preservatives, stabilisers, buffers, antioxidants and/or other
additives may be included, as required.

[0137] Whether it is a polypeptide, antibody, peptide, nucleic acid molecule,
small molecule or other pharmaceutically useful compound that is to be given to
an individual, administration is preferably in a "prophylactically effective
amount" or a "therapeutically effective amount" (as the case may be, although
prophylaxis may be considered therapy), this being sufficient to show benefit
to the individual. The actual amount administered, and rate and tir[alpha]e-
course of administration, will depend on the nature and severity of what is
being treated. Prescription of treatment, e.g. decisions on dosage etc, is
within the responsibility of general practitioners and other medical doctors,
and typically takes account of the disorder to be treated, the condition of the
individual patient, the site of delivery, the method of administration and other
factors known to practitioners. Examples of the techniques and protocols
mentioned above can be found in Remington' s Pharmaceutical Sciences, 20th
Edition, 2000, pub. Lippincott, Williams & Wilkins.

[0138] Alternatively, targeting therapies may be used to deliver the active
agent more specifically to certain types of cell, by the use of targeting
systems such as antibody or cell specific ligands. Targeting may be desirable
for a variety of reasons; for example if the agent is unacceptably toxic, or if
it would otherwise require too high a dosage, or if it would not otherwise be
able to enter the target cells.

[0139] A composition may be administered alone or in combination with other
treatments, either simultaneously or sequentially dependent upon the condition
to be treated.

[0140] Examples A group at the University of UIm, Germany, has recently
suggested that a pentapeptide (QYNAD) with Na<+> channel-blocking function could
be a biological marker of certain inflammatory and immunological disorders of
the nervous system<16>.

[0141] The inventors asked whether or not the pentapeptide identified by the
German group<16> might play a role in CFS. Samples of serum were sent to the
University of UIm for analysis. The 15 samples included 5 normal controls, 5
patients with CFS and 5 disease controls including two patients with MS.
Samples were numbered 1-15 and the German group were not informed what the
samples were, or which samples were which, until the experiment was concluded.

[0142] When the code was broken, the results showed that the disease control
group had levels of the pentapeptide which were 2.3X those of the normal
controls (similar to the published data) and the CFS samples had levels which
were 3X higher than the healthy controls.

[0143] Thus, there are measurably higher amounts of the pentapeptide in
patients with CFS compared with healthy controls. Although the pentapeptide may
not be specific to CFS (as high levels are also found in other disorders), an
assay for the peptide could be used as part of the differential diagnosis of CFS.

[0144] The German group was unable to identify an endogenous gene which encodes
the pentapeptide. The inventors carried out NCBI BLAST and EMBL-Heidelberg
Bioccelerator amino acid alignments for the pentapeptide QYNAD. A total of one
hundred alignment hits were found. Of these, only nine showed 100% similarity
over the five amino acids - five of those were human. The amino acid searches
were followed by NCBI BLAST searches using the GenBank Accession and gi numbers
for each of the five human amino acid to determine their origins, references
and nucleotide sequences. A number of cloned nucleotide sequences were found
and when these were run through the nucleotide databases, only one clone showed
full-length homology to any human gene. This gene was a human ion-channel gene
- the vacuolar proton pump H<+>-ATPase (v-ATPase). Remarkably, when the human
gene amino acid sequence was compared with the original QYNAD pentapeptide it
was discovered that the relevant part of the human ion channel encodes the
sequence QYMAD.

[0145] The inventors next asked whether the v-ATPase represents a candidate gene
for a diagnostic test for CFS. RT-PCR using primers specific for the v-ATPase
was performed on cDNA prepared from mRNA from PBMCs from CFS patients and
healthy controls. As shown in Figure 1, the patient samples have a
significantly higher level of v-ATPase mRNA than the healthy controls. Thus the
v-ATPase gene appears to represent a genuine biomarker for CFS/ME.

[0146] The vATPase is known to be involved in regulation of a number of
metabolic functions which are deranged in CFS/ME. vATPase upregulation could
therefore provide an explanation for a number of the symptoms observed. For
example, increased vATPase activity could explain the intracellular acidosis in
exercising muscles, chest pain (syndrome X), altered neurotransmitter
(dopamine) function and abnormal regulation of hypothalamic hormones. In
addition, it could explain the increased energy expenditure and fatigue
associated with the condition. Taken together, this suggests that the vATPase is
not only a marker for the condition, but is a realistic target for intervention
therapy.

[0147] Other biomarkers for CFS/ME

[0148] The inventors went on to examine whether the increase in vATPase
expression was confirmed by microarray analysis. Such analysis provides the
opportunity to examine the differential expression of mRNA from a very large
number of genes. Surprisingly, the results of the analysis not only confirmed
their earlier findings regarding the vATPase gene, but also identified
differences in the level of expression of key genes in the PBMC of patients with
CFS/ME and control subj ects, giving an insight into the biochemical pathways
which are involved in this disorder.

[0149] Microarray results have been verified by western blot analysis and RT-PCR
assay. A number of genes, in addition to v-ATPase, were significantly
up/downregulated and identified as suitable biomarkers for the disorder.

[0150] Advances in genome sequencing and automated chip manufacture have made
DNA chip or microarray technology readily available<25>. This technology allows
simultaneous differential expression profiling from a very large number of genes
in tissue samples of CFS/ME patients and controls. A recent report from Vernon
et al (2002), described a CFS biomarker search in PBMC using a DNA chip array
assay which included 1, 764 genes. In the study reported here, RNA isolated
from PBMC, was assayed using Affymetrix genome-wide chips (HG-U133 arrays)
which included 30,000 gene sequences.

[0151] Using DNA microarray analysis of whole human genome, gene
transcriptional signatures were compared in the PBMC of eight male patients with
CFS and seven age-matched male healthy controls. An additional cohort of
fourteen patients with CFS and age and sex matched controls was recruited for
RT-PCR and western blot assays in order to verify the microarray data. Analysis
of the microarray data was performed as described previously (Breitling R.
Armengaud P. Amtmann A. Herzyk P. Rank products: A simple, yet powerful, new
method to detect differentially regulated genes in replicated microarray
experiments. FEBS Letters 2004; 573 (1-3): 83-92; Breitling R. Amtmann A.
Herzyk P. Iterative Group Analysis (iGA): A simple tool to enhance sensitivity
and facilitate interpretation of microarray experiments. BMC Bioinformatics
2004; 5: (pp 8p)).

[0152] Genes which are significantly upregulated in CFS/ME patients compared to
healthy controls are detailed in Table 6, ranked according to their RP values.
It is considered that any of these genes may be used as biomarkers for CFS/ME.
Particularly preferred marker genes are detailed in Table 1.

[0153] Further genes, including prostaglandin D2 synthase and T-cell receptors
alpha, beta, gamma and delta are found to be downregulated in CFS patients
compared to normal controls. Prostaglandin D2 synthase (NCBI accession no. BC
005939, UniGene Hs. 446429) is considered to be a good candidate for a CFS/ME
biomarker, because it is known to be involved in sleep regulation; patients with
CFS frequently suffer from sleep reversal and fatigue associated with lack of
sleep. However, data for other downregulated genes is not shown here. In
general, genes which are upregulated in the disease state are considered to be
better biomarkers for diagnostic tests etc. than genes which are downregulated
because the potential for false-positive tests is significantly higher when
using genes which are underexpressed in the disease state.

[0154] Iterative group analysis of the differentially expressed genes indicate
that in CFS, there is a shift of immune response with preferential antigen
presentation to MHC class Il receptors and downregulation of T-cell
receptor-[alpha], increased cell membrane prostaglandin-endoperoxide synthase
activity with downstream changes in oxygen transport and also activation of the
guanyl cyclase and caspase pathways of cellular apoptosis. Another set of genes
was identified which are involved in the immediate response to infection,
particularly by intracellular parasites. The particular genes involved in each
of these pathways are identified in Tables 2 to 5. In each of these key pathways,
the hub genes were higher ranked in the analysis compared to the network genes.

[0155] Functional changes produced by altered gene regulation may explain the
mechanism of fatigue and offer a rational basis for targeted pharmacotherapy in
CFS.

[0156] Table 1 - Biomarker Genes for CFS/ME

[0157]
<EMI ID=36.1>


[0158]
<EMI ID=37.1>

[0159] Table 2: Genes involved in defence against infection by intracellular
pathogens

[0160]
<EMI ID=37.2>

[0161]
<EMI ID=38.1>

[0162] Table 3: Genes involved in oxidative stress response

[0163]
<EMI ID=38.2>

[0164] Table 4: Genes involved in apoptosis

[0165]
<EMI ID=38.3>

[0166] Table 5: Genes involved in antigen presentation and immunity

[0167]
<EMI ID=38.4>

[0168]
<EMI ID=39.1>

[0169] NCBI accession numbers refer to the UniGene database, build no. 177,
released 22 December 2004.

[0170] It is clear from this data that significant differences in the expression
of a number of genes can be seen in PBMC samples from patients with CFS and
healthy controls. We have verified that the DNA microarray assay is valid by
confirming the results by RT-PCR and western blot analyses. This is the first
time that a reproducible biochemical lesion has been seen in patients with CFS.
We propose that bioassays of the significantly over-expressed genes could be
used as diagnostic biomarkers for CFS to aid in the differential diagnosis of
the condition.

[0171] In order to confirm the relevance of the genes identified in the
microarray experiments, Western blot and RT-PCR assays have been performed to
analyse the expression of selected genes in samples from patients and controls
different to those studied in the microarray analysis. The results verify that
these genes may be used as potential biomarkers to support the clinical
diagnosis of CFS and identify suitable candidates for treatment trials.

[0172] Figures 2 to 5 show upregulation of mRNA and/or proteins from various of
these genes in PBMCs from patients suffering from CFS/ME, as compared to age-
and sex-matched healthy controls. Figure 2 shows RT-PCR analysis of Defensin
alpha 1, Figures 3 and 4 show Western blot analysis of Defensin alpha 1 and
Thrombospondin 1 respectively. In all cases, there is significant upregulation
of the protein or mRNA in the disease group compared to the control group.
Figure 6 shows a duplicate of the SDS-PAGE gel used for the Western blot,
stained with Coomassie blue. This confirms that the observed differences in
protein levels between the disease and control groups are not due to unequal
loading of total protein on the gels.

[0173] Discussion

[0174] Previous reports have hypothesised that CFS is a form of channelopathy -
a disorder of membrane ion channels<9>' <11>' <13>. There are several reports in
the literature which we believe strengthen the hypothesis that the vacuolar H+
ATPase plays a pathogenic role in CFS.

[0175] Local anaesthetics, which are known to act on ion channels, have an
adverse effect on patients with CFS/ME. It has been demonstrated also, that in
some patients with CFS/ME, there are morphological changes to the red blood
cells<19>. Remarkably, a study by Nishiguchi et al<20>, has demonstrated that
the local anaesthetic lidocaine can induce reversible morphological
transformation of human red blood cells and that this change is mediated by the
activation of vacuolar H+ ATPase. In addition, Li et al<21a> have shown that
the gene is involved in iron binding in red blood cells.

[0176] The ion channel gene is a member of the vacuolar H+ ATPase proton
transporting gene family<21>'<22>'<23>. This family of genes is directly
involved with the phosphocreatine-dependent glutamate uptake by synaptic
vesicles<24>. The gene is responsible for vesicle docking/exocytosis during
neurotransmiter release<25> and is a major constituent of synaptic vesicles
associated with intracellular membrane structures<26>. We have demonstrated,
using <1>H MRS that there is a perturbation of the choline/creatine balance in
the CNS (Condon et al<17>, Chaudhuri et al<42>). This finding has been
corroborated by Puri et al<18>. As stated above, this type of gene is directly
involved in the creatine pathways. We have previously demonstrated that
patients with CFS have low body- potassium levels<9>. Bailey et al<27> have
shown a relationship between potassium depletion and up-regulation of H+-ATPase.

[0177] As stated above, viruses have often been associated with CFS. Virus
entry into cells may be mediated by H+ATPase<28>'<29>'<30>. In addition to viral
infection affecting neurotransmitter function<7>, there is a large body of
evidence to show that the vacuolar H+-ATPase is also invoked<31>'<32>'<33,34>'
<35>'<36>'<37>'<38, 39.>

[0178] It is clear from the above data that significant differences in the
expression of a number of genes can be seen in PBMC samples from patients with
CFS and healthy controls. This is the first time that a reproducible
biochemical lesion has been seen in patients with CFS We propose that bioassays
of the significantly over-expressed genes (below) could be used as diagnostic
biomarkers for CFS to aid in the differential diagnosis of the condition.

[0179] Materials and Methods

[0180] Study Subjects

[0181] Patients with CFS were diagnosed with reference to the 1994 Fukuda
definition. All seven patients were male, aged between 18 and 54 years (mean
36), and were not on medication. Healthy control subj ects were male, aged
between 22 and 58 years (mean 34). In addition to the 8 patients and seven
control subjects used for the DNA chip assays, an additional fourteen patients
and controls were used to confirm the chip assay results by RT-PCR and western
blot assays. Informed consent and ethical approval were obtained.

[0182] Isolation of peripheral blood mononuclear cells (PBMC) for protein
purification

[0183] Venous blood samples were drawn from patients who fulfilled the

[0184] Holmes and Fukuda criteria for ME/CFS, and from healthy individuals. The
procedure for isolating PBMC was started immediately and finished within 2h of
sampling. EDTA treated whole blood was diluted 1: 1 with phosphate buffered
saline. Two volumes of blood were overlaid on one volume of Histopaque -1077
(Sigma Diagnostics) and centrifuged at 20[deg.]C AT 50Og for 30 min. The PBMC
interface was removed and washed twice with phosphate buffered saline and
centrifuged. The pellets were resuspended in phosphate buffered saline, an
aliquot removed and counted in red blood cell lysis buffer (155Mm NH4CL, 10Mm
NaHCO3, pH7.4, 0.1mM EDTA). The PBMC were centrifuged once more (20[deg.]C,
500g, 10min). The PBMC were then aliquoted into micortubes equivalent to 5x105
cells per tube, centrifuged and stored as dry pellets at -80[deg.]C.

[0185] Western Blot Analysis

[0186] PBMC pellets were resuspended in sample reducing buffer (ImI Glycerol,
0.5ml [beta]-mercaptoethanol, 3ml 10% SDS, 1.25ml IM Tris-HCL Ph 6.7), boiled
for 5 min. The lysates were loaded onto a 10% PAGE gel, each track equivalent
to 2x 10 5 PBMC. The PAGE gel was assessed for equal protein load by coomassie
stain. The gel then electrophoretically transferred onto nitrocellulose PVDF
membrane (Biorad) for 2 hours. The blots were blocked for non specific binding
with 10% normal goat serum for 30 min, probed with a mouse Mab to human
defensinl-3, (Hycult, Netherlands), mouse Mab to human thrombospondin
(Sigma-Aldrich Inc) and a mouse Mab to Chondroitin Sulphate Proteoglycan
(USBiological MA USA), at dilutions of 1/100, 1/1000, 1/1000 respectively in
TBS 0.05% Tween 20 for 2 hours at RT. The protein was detected after subsequent
incubation with alkaline phosphatase conjugate Goat anti Mouse IgG (1/1000
final dilution) (Jackson Immunoresearch Laboratories PA USA). The reactions
were detected using SIGMA FAST BCIP/NBT).

[0187] Development of the PCR assay for the CFS gene.

[0188] Oligonucleotide primers which span a specific epitope within the CFS gene
were chosen and tested by RT-PCR. RNA from blood samples from patients with CFS
and appropriate controls were RT-PCR amplified and PCR amplicons quantitated by
gel documentation system software.

[0189] RNA Isola tion for RT-PCR

[0190] Total RNA was isolated from peripheral blood mononuclear cells (PBMC)
using the Promega RNAgents Total RNA Isolation System.

[0191] Venous blood was collected in standard EDTA blood tubes and RNA purified
using the method of Chomczynski and Sacchi. White blood cell pellets were
homogenised by hand in an appropriate volume of denaturing solution (guanidinium
thiocyanate, 4M; sodium citrate, 25mM; N-laurolyl sarcosine, 0.5%;
2-mercaptoethanol, 0.1M, in distilled water adjusted to pH 7.0). To homogenate
(ImI), sodium acetate (100[mu]l, 2M, pH 4.0), citrate-buffered (0.1M, pH
4.3) phenol

[0192] { lml) and chloroform: isoamyl alcohol (49: 1, 200[mu]l) were
sequentially added. The resulting mixture was treated in a vortex mixer (Fisons
Scientific Equipment, Whirlimixer(TM)) for 10 seconds then incubated on ice for
15 minutes. Samples were then centrifuged (12, 000g, 20 minutes, 4[deg.]C) and
the upper aqueous layer pippetted into to a fresh tube. After addition of ice
cold isopropanol (lml), RNA precipitated from the mixture during a thirty
minute period on dry ice. The mixtures were then centrifuged (12, 000g, 20
minutes, 4[deg.] C) and the supernatant discarded. The pellet was dissolved in
denaturing solution (300[mu]l) and transferred to a microcentrifuge tube (
1.5ml, Axygen). Ethanol (absolute, 600[mu]l) was added to each microcentrifuge
tube, samples were incubated on dry ice for 30 minutes, and then centrifuged (
1160Og, 20 minutes, 4[deg.]C). The resulting pellet were then washed twice in
ethanol (70% aqueous solution) then lyophylised (using a Hetosicc freeze-drier
and a JAVAC high vacuum pump DD-75). The freeze-dried RNA was re-suspended in
sterile distilled water (60[mu]l) and stored at -70[deg.]C until required.

[0193] Quantifica tion and examination of RNA

[0194] RNA solution (prepared as described above, 2[mu]l) was added to distilled
water (98[mu]l) to produce a 1: 50 dilution. The optical density of the
sample was read at 260nn[alpha] and 280nm using a 50[mu]l ultraviolet cuvet.
The absorbance ratio 260/280nm measured at these wavelengths indicates the
purity of the RNA. The absolute concentration of RNA was estimated using the
following equation:

[0195] Optical Density260nm x 40 x dilution = RNA concentration ([mu]g/ml)

[0196] To examine the quality, 5[mu]l of RNA was added to 5[mu]l of the gel
marker Orange G (1% Orange G dye in 50% glycerol, 50% 2x TBE). This was heated
to 70[deg.]C for 3 minutes, then electrophoresed through a horizontal 1%
agarose gel in 1 x TBE (10x TBE = 0.089M tris (hydroxymethyl) - methylamine,
0.089M boric acid, 0.025M disodium EDTA, pH 8.3). The gel was then stained for
30 minutes in ethidium bromide (0.5 mg/ml) in TBE, de-stained in water and
visualised under medium wave (320nm) ultraviolet light.

[0197] cDNA synthesis

[0198] Two micrograms of RNA were added to 1[mu]l oligo (dT) 12-18 (
500[mu]g/ml, Roche) and the volume made up to 11[mu]l with sterile distilled
water.

[0199] This was incubated (70[deg.]C, 10 minutes) to allow the oligo (dT) to
bind to the poly-A tail of the RNA, and then chilled on ice. To the reaction
mixture, 4[mu]l of 5x First Strand Buffer, 2[mu]l of 0. IM DTT, 1[mu] l of 10mM
dNTP mix (10mM each dATP, dGTP, dCTP, dTTP; Amersham Pharmacia Biotech),
1[mu]l of sterile distilled water and 1[mu]l of

[0200] Superscript II (Gibco-BRL(R) Life Technologies) were added. This was
incubated first at 50[deg.]C for Ihr, then 70[deg.]C for 15 minutes to
inactivate the reaction. A negative cDNA control (2[mu]l of distilled water)
was included in each cDNA synthesis to confirm that the reaction mix was not
contaminated.

[0201] Polymerase Chain Reaction To 2[mu]l of cDNA, 10[mu]l of 10x
magnesium-free buffer, 10[mu]l of 2.5mM dNTPs (2.5mM each dATP, dGTP, dCTP,
dTTP), 6[mu]l MgCl2 (25mM) and 1[mu]l each of the appropriate 5' and 3 '
primers (0.5[mu]g/[mu]l) were added. The volume was made up to 99.8[mu]l using
sterile distilled water and 0.2[mu]l of Taq DNA polymerase (Promega) was added.
The PCR reaction (35 cycles) was carried out on a Techne Genius thermocycler.

[0202] Primers used to amplify vATPase were 5 ' -etc gtg ace tgt tac tgc tg - 3
' and 5 ' -aag taa cca agt cca etc ca-3 '. Primers for Defensin 1 were 5 ' -caa
gag ctg atg agg ttg ct-3 ' and 5 ' -gaa ggt aca gga gta ata gc-3 '.

[0203] Thirty microlitres of PCR product was added to 5[mu]l of orange G and
electrophoresed through a horizontal 2-3% agarose gel in 1 x TBE at 100 volts,
until the dye front had migrated a minimum of 10cm. On each gel, 3[mu]l of a 123
base pair DNA ladder in 5[mu]l of Orange G was included as a size marker for
comparison with PCR product bands. The gel was then stained for 30 minutes in
ethidium bromide (0.5 mg/ml) in TBE, destained in water and visualised under
medium wave (320nm) ultraviolet light.

[0204] Gene expression was analysed by measuring the band density for each
amplicon. Band densities were measured using the Herolab EASY Plus computer
automated image analysis system. True comparisons in gene expression between
samples were enabled by comparing results of densitometry for the experimental
genes against the housekeeping gene (abl - tyrosine kinase) band densities.
References

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Ishizuka, S., Nakayama, K., Yanai, M., Numazaki, Y., Sasaki, H.
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[0232] 34. Moriyama, Y., Tsai, H. L. & Futai, M. Energy-dependent
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[0240] 42. Chaudhuri A, Gow JW Behan PO, "Chronic fatigue syndrome and systemic
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[0241] 43. Vernon, SD, Unger, ER, Dimulescu, IM, Rajeevan, M and Reeves, WC.
Utility of the blood for gene expression profiling and biomarker discovery in
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Salvato FR, Morgan R, Fletcher MA. Immunological abnormalities in chronic
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[0242] 45. Lloyd AR, Wakefield D, Hickie I. Immunity and the pathophysiology of
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[0243] 46. Barker E, Fuj imara SF, Fadem MB, Landay AL, Levy JA. Immunological
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[0245] 48. Vollmer-Conna U, Lloyd A, Hickie I, Wakefield D. Chronic fatigue
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Table 6

[0246]
<EMI ID=52.1>

[0247]
<EMI ID=52.2>

[0248]
<EMI ID=53.1>

[0249]
<EMI ID=54.1>

[0250]
<EMI ID=55.1>

[0251]
<EMI ID=56.1>

[0252]
<EMI ID=57.1>

[0253]
<EMI ID=58.1>

[0254]
<EMI ID=59.1>

[0255]
<EMI ID=60.1>

[0256]
<EMI ID=61.1>

[0257]
<EMI ID=62.1>

[0258]
<EMI ID=63.1>

[0259]
<EMI ID=64.1>

[0260]
<EMI ID=65.1>

[0261]
<EMI ID=66.1>

[0262]
<EMI ID=67.1>

[0263]
<EMI ID=68.1>

[0264]
<EMI ID=69.1>

[0265]
<EMI ID=70.1>

[0266]
<EMI ID=71.1>

[0267]
<EMI ID=72.1>

[0268]
<EMI ID=73.1>

[0269]
<EMI ID=74.1>

[0270]
<EMI ID=75.1>

[0271]
<EMI ID=76.1>

[0272]
<EMI ID=77.1>

[0273]
<EMI ID=78.1>

[0274]
<EMI ID=79.1>

[0275]
<EMI ID=80.1>

[0276]
<EMI ID=81.1>

[0277]
<EMI ID=82.1>

[0278]
<EMI ID=83.1>

[0279]
<EMI ID=84.1>

[0280]
<EMI ID=85.1>

[0281]
<EMI ID=86.1>

[0282]
<EMI ID=87.1>

[0283]
<EMI ID=88.1>

[0284]
<EMI ID=89.1>

[0285]
<EMI ID=90.1>

[0286]
<EMI ID=91.1>

[0287]
<EMI ID=92.1>

[0288]
<EMI ID=93.1>

[0289]
<EMI ID=94.1>

[0290]
<EMI ID=95.1>

[0291]
<EMI ID=96.1>

[0292]
<EMI ID=97.1>

[0293]
<EMI ID=98.1>

[0294]
<EMI ID=99.1>

[0295]
<EMI ID=100.1>

[0296]
<EMI ID=101.1>

[0297]
<EMI ID=102.1>

[0298]
<EMI ID=103.1>

[0299]
<EMI ID=104.1>

[0300]
<EMI ID=105.1>

[0301]
<EMI ID=106.1>

[0302]
<EMI ID=107.1>

[0303]
<EMI ID=108.1>

[0304]
<EMI ID=109.1>

[0305]
<EMI ID=110.1>

[0306]
<EMI ID=111.1>

[0307]
<EMI ID=112.1>

[0308]
<EMI ID=113.1>

[0309]
<EMI ID=114.1>

[0310]
<EMI ID=115.1>

[0311]
<EMI ID=116.1>

[0312]
<EMI ID=117.1>

[0313]
<EMI ID=118.1>

[0314]
<EMI ID=119.1>

[0315]
<EMI ID=120.1>

[0316]
<EMI ID=121.1>

[0317]
<EMI ID=122.1>

[0318]
<EMI ID=123.1>

[0319]
<EMI ID=124.1>

[0320]
<EMI ID=125.1>

[0321]
<EMI ID=126.1>

[0322]
<EMI ID=127.1>

[0323]
<EMI ID=128.1>

[0324]
<EMI ID=129.1>

[0325]
<EMI ID=130.1>

[0326]
<EMI ID=131.1>

[0327]
<EMI ID=132.1>

[0328]
<EMI ID=133.1>

[0329]
<EMI ID=134.1>

[0330]
<EMI ID=135.1>

[0331]
<EMI ID=136.1>

[0332]
<EMI ID=137.1>

[0333]
<EMI ID=138.1>

[0334]
<EMI ID=139.1>

[0335]
<EMI ID=140.1>

[0336]
<EMI ID=141.1>

[0337]
<EMI ID=142.1>

[0338]
<EMI ID=143.1>

[0339]
<EMI ID=144.1>

[0340]
<EMI ID=145.1>

[0341]
<EMI ID=146.1>

[0342]
<EMI ID=147.1>

[0343]
<EMI ID=148.1>


Claims of WO2006082390
----------------------

Claims

1. A method for investigating whether a test subj ect is affected by
chronic fatigue syndrome/myalgic encephalomyelitis (CFS/ME), comprising
providing a biological sample from the subj ect and determining the level
in the sample of one or more biomarkers for CFS/ME, wherein the biomarkers
comprise an expression product of at least one gene shown in Table 1.

2. A method according to claim 1, wherein the biomarkers comprise
expression products of a plurality of genes shown in Table 1.

3. A method according to claim 2 wherein the biomarkers comprise
expression products of 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20 or more
of the genes shown in Table 1.

4. A method according to any one of claims 1 to 3 wherein the biomarkers
comprise expression products of one or more of defensin [alpha]1,
haemoglobin [gamma], CXCR4, tubulin beta 1 and HLA DR[beta]4 genes.

5. A method according, to claim 4 wherein the biomarkers comprise
expression products of two, three, four or five of defensin [alpha]1,
haemoglobin [gamma], CXCR4, tubulin beta 1 and HLA DR[beta]4 genes.

6. A method according to claim 4 or claim 5 wherein the biomarkers
comprise expression products of further hub genes of Table IA.

7. A method according to any one of claims 4 to 6 wherein the biomarkers
further comprise expression products of one or more of thrombospondin 1,
caspase 1 and/or IgG3.

8. A method according to any one of claims 1 to 7 wherein the biomarkers
comprise expression products of at least one gene from each of at least
two of Tables 2, 3, 4 and 5.

9. A method according to claim 8 wherein said at least one gene from each
said Table is a hub gene.

10. A method according to any one of the preceding claims wherein the
biomarkers comprise expression products of defensin cxl, CXCR4 and
lactotransferrin (LTF) genes.

11. A method according to claim 10 wherein the biomarkers comprise
expression products of defensin [alpha]4, integrin [alpha]2B, integrin
[beta]3, arginase 1, arginase 2, thrombospondin 1, membrane associated
protein 17 (MAP 17 ), Charcot Leyden Crystal Protein (CLC) and/or
chondroitin sulphate proteoglycan 2 (versican) genes.

12. A method according to any one of the preceding claims wherein the
biomarkers comprise an expression product of haemoglobin [gamma] (foetal
haemoglobin).

13. A method according to claim 12 wherein the biomarkers comprise
expression products of haemoglobin alpha 1, haemoglobin beta, haemoglobin
delta, prostaglandin-endoperoxide synthase 1 and/or
prostaglandin-endoperoxide synthase 2 genes.

14. A method according to any one of the preceding claims wherein the
biomarkers comprise an expression product of serine/threonine kinase 17B
(STK17B).

15. A method according to claim 14 wherein the biomarkers comprise
expression products of caspase 1, dynamin 1-like, and/or phosphatidyl
serine binding protein genes.

16. A method according to any one of the preceding claims wherein the
biomarkers comprise an expression product of HLA-DR[beta]4.

17. A method according to claim 16 wherein the biomarkers comprise
expression products of HLA-DQ[beta]l and/or immunoglobulin heavy chain
[gamma]3 genes.

18. A method according to any one of the preceding claims wherein the
biomarkers comprise an expression product of one or more further genes
from Table 6.

19. A method according to any one of the preceding claims wherein the
biomarkers comprise an expression product of prostaglandin D2 synthase.

20. A method for classifying a subject affected by CFS/ME, the method
comprising providing a biological sample from the subj ect and determining
the level in the sample of one or more biomarkers for CFS/ME, wherein the
biomarkers comprise expression products of at least one gene shown in one
or more of Tables 2 to 5.

21. A method according to claim 20 wherein the biomarkers comprise
expression products of a hub gene from one of Tables 2, 3, 4 or 5.

22. A method according to claim 21 wherein the biomarkers comprise
expression products of hub genes from each of two, three or all of Tables
2, 3, 4 or 5.

23. A method according to any one of claims 20 to 22 further comprising
determining network genes from one, two, three or all of Tables 2, 3, 4
and 5.

24. A method according to any one of claims 20 to 23 wherein the
biomarkers comprise expression products of defensin [alpha]1, CXCR4 and
lactotransferrin (LTF) genes.

25. A method according to claim 24 wherein the biomarkers comprise
expression products of defensin [alpha]4, integrin [alpha]2B, integrin
[beta]3, arginase 1, arg+-nase 2, thrombospondin 1, membrane associated
protein 17 (MAP 17 ), Charcot Leyden Crystal Protein (CLC) and/or
chondroitin sulphate proteoglycan 2 (versican) genes.

26. A method according to any one of claims 20 to 25 wherein the
biomarkers comprise an expression product of haemoglobin Y ( foetal
haemoglobin).

27. A method according to claim 26 wherein the biomarkers comprise
expression products of haemoglobin alpha 1, haemoglobin beta, haemoglobin
delta, prostaglandin-endoperoxide synthase 1 and/or
prostaglandin-endoperoxide synthase 2 genes.

28. A method according to any one of claims 20 to 27 wherein the
biomarkers comprise an expression product of serine/threonine kinase 17B
(STK17B).

29. A method according to claim 28 wherein the biomarkers comprise
expression products of caspase 1, dynamin 1-like, and/or phosphatidyl
serine binding protein genes.

30. A method according to any one of claims 20 to 28 wherein the
biomarkers comprise an expression product of HLA-DR[beta]4.

31. A method according to claim 30 wherein the biomarkers comprise
expression products of HLA-DQ[beta]l and/or immunoglobulin heavy chain
[gamma]3 genes.

32. A method according to any one of claims 20 to 31 comprising
classifying the subject as being affected by one or more of oxidative
stress, excessive apoptosis, and immunological dysregulation (MHC I to II
shift).

33. A method of determining whether an individual affected by CFS/ME is
suitable for treatment with an anti-oxidant, minocycline or a
corticosteroid, comprising determining whether the individual is affected
by one or more of oxidative stress, excessive apoptosis, and immunological
dysregulation by a method according to any one of claims 20 to 24, and
prescribing a suitable treatment depending on the outcome.

34. A method of creating a library of expression profiles for use in
determining whether an individual is affected by CFS/ME, the method
comprising
(a) providing biological samples from a plurality of individuals affected
by CFS/ME, and determining the level in each sample of one or more
biomarkers for CFS/ME to create a plurality of expression profiles from
affected individuals;
(b) providing biological samples from a plurality of individuals not
affected by CFS/ME, and determining the level in the sample of said one or
more biomarkers for CFS/ME to create a plurality of expression profiles
from unaffected individuals;
wherein the biomarkers comprise expression products of one or more genes
shown in Table 1.

35. A method according to claim 34 comprising the step of retrievably
storing each of the expression profiles on a computer data carrier, in
order to create a database of expression profiles for both affected and
unaffected individuals.

36. A method of determining the efficacy of a treatment for CFS/ME
comprising the steps of:
(a) providing a biological sample from a subj ect affected by CFS/ME who
has been subj ected to said treatment, (b) determining the level in said
sample of one or more biomarkers for CFS/ME to create an expression
profile for said subj ect, and
[c) comparing said expression profile with
i) a comparable expression profile obtained from said test subject before
initiation of said treatment, and/or
ii) a comparable expression profile obtained from said test subject at an
earlier stage of said treatment, and/or
iii) a comparable expression profile characteristic of a subj ect who is
unaffected by CFS/ME,
wherein the one or more biomarkers for CFS/ME comprise expression products
of one or more genes shown in Tables 1 to 5.

37. A method according to any one of the preceding claims wherein the
biological sample is a blood sample or comprises peripheral blood
mononuclear cells isolated from a blood sample.

38. A method according to any one of the preceding claims wherein the
expression level of each of the biomarkers is determined by contacting the
sample with a binding agent capable of binding specifically to said
expression product of the corresponding gene.

39. A method according to claim 38 wherein the expression product is a
nucleic acid.

40. A method according to claim 39 wherein the binding agent is a nucleic
acid probe or primer.

41. A method according to claim 39 or claim 40 wherein the expression
product is mRNA, pre-r[alpha]RNA or cDNA.

42. A method according to any one of claims 39 to 41 wherein the
expression level of the biomarker is determined by means of Northern blot,
in situ hybridisation, RT-PCR, Southern blot or microarray assay.

43. A method according to claim 38 wherein the expression product is a
polypeptide.

44. A method according to claim 43 wherein the binding agent is a ligand
or receptor for said polypeptide, or an antibody or fragment thereof
specific for said polypeptide

45. A method according to claim 43 or claim 44 wherein the expression
level of the biomarker is determined by Western blot, ELISA or microarray
assay.

46. A method according to any one of the preceding claims further
comprising the step of determining the level, in the biological sample,
or in a further biological sample from the subj ect, of the peptide QYNAD.

47. A method according to claim 46 wherein the further biological sample
is a blood or serum sample.

48. A method according to claim 46 wherein the further biological sample
is cerebrospinal fluid.

49. A kit for use in a method according to any one of the preceding claims,
the kit comprising a plurality of binding agents, each capable of
binding specifically and individually to an expression product of one of
the genes of Table 1, or the peptide QYNAD.

50. A kit according to claim 49 comprising binding agents specific for
expression products of two or more genes of Tables 1, or at least one
gene of Table 1 and the peptide QYNAD.

51. A kit according to claim 50 comprising binding agents capable of
binding specifically and individually to expression products of 2, 3, 4,
5, 6, 7, 8, 9, 10, 15, 20 or more of the genes of Table 1 and
optionally the peptide QYNAD.

52. A kit according to any one of claims 49 to 51 wherein the binding
agents are immobilised on one or more solid supports.

53. A kit according to any one of claims 49 to 52 further comprising one
or more binding agents capable of binding specifically to an expression
product of a control gene which is substantially not differentially
expressed between individuals affected and whose expression level is
substantially unaffected by CFS/ME.

54. A kit according to claim 53 wherein the control gene is glyceraldehyde
phosphate dehydrogenase (GAPDH), [beta]-actin, or abl (ableson tyrosine
kinase).

55. A kit according to any one of claims 49 to 54 comprising binding
agents for expression products of less than 1000 different genes, less
than 500 different genes, less than 100, less than 50, less than 40,
less than 30, less than 20, or less than 10 different genes.

56. A kit according to any one of claims 49 to 55 wherein the expression
product is a nucleic acid and the binding agent is a nucleic acid probe or
primer.

57. A kit according to claim 56 wherein the expression product is mRNA,
pre-mRNA or cDNA.

58. A kit according to any one of claims 49 to 55 wherein the expression
product is a polypeptide and the binding agent is a ligand or receptor for
said polypeptide, or an antibody or fragment thereof specific for said
polypeptide

59. An expression profile database, comprising a plurality of expression
profiles of biomarkers for CFS/ME from affected and unaffected individuals
, wherein the biomarkers comprise expression products of one or more genes
shown in Table 1.

60. A method of treating CFS/ME in an individual suffering therefrom,
comprising administering an effective amount of an antioxidant.

61. A method according to claim 60 further comprising administering
therapeutically effective amounts of a corticosteroid and/or minocyline.

62. Use of an anti-oxidant in the preparation of a medicament for the
treatment of CFS/ME.

63. Use according to claim 62 wherein the medicament is formulated for
administration in conjunction with a corticosteroid and/or minocycline, or
comprises a corticosteroid and/or minocycline.

64. A method according to claim 60 or claim 61, or use according to claim
62 or claim 63, wherein the anti-oxidant is coenzyme Q10 or an inhibitor
of a cyclooxygenase (COX) enzyme such as celecoxib (4- [5-
(4-methylphenyl) -3- (trifluoromethyl ) -1H-pyrazol-1-yl]
benzenesulfonamide).

65. A method of treating CFS/ME in an individual suffering therefrom,
comprising administering an effective amount of minocycline.

66. A method according to claim 65 further comprising administering a
therapeutically effective amount of a corticosteroid and/or an
anti-oxidant.

67. Use of minocycline in the preparation of a medicament for the
treatment of CFS/ME.

68. Use according to claim 67 wherein the medicament is formulated for
administration in conjunction with a corticosteroid and/or an
anti-oxidant, or comprises a corticosteroid and/or an anti-oxidant.

69. A method of treating CFS/ME in an individual suffering therefrom,
comprising administering an effective amount of a corticosteroid.

70. A method according to claim 69 further comprising administering a
therapeutically effective amount of minocycline and/or an anti-oxidant.

71. Use of a corticosteroid in the preparation of a medicament for the
treatment of CFS/ME.

72. Use according to claim 71 wherein the medicament is formulated for
administration in conjunction with minocycline and/or an antioxidant, or
comprises minocycline and/or an anti-oxidant.

73. A method according to claim 69 or claim 70, or use according to claim
71 or claim 72, wherein the corticosteroid is hydrocortisone,
dexamethasone or prednisone.

74. A pharmaceutical composition comprising a therapeutically effective
amount of minocycline in combination with a therapeutically effective
amount of a corticosteroid and/or an anti- oxidant, and a pharmaceutically
acceptable carrier..

75. A pharmaceutical composition comprising a therapeutically effective
amount of a corticosteroid in combination with a therapeutically effective
amount of minocycline and/or an anti- oxidant, and a pharmaceutically
acceptable carrier.

76. A pharmaceutical composition comprising a therapeutically effective
amount of an anti-oxidant in combination with a therapeutically effective
amount of a corticosteroid and/or minocycline, and a pharmaceutically
acceptable carrier.


Mosaics of WO2006082390
-----------------------

[See European Patent Office Document
     http://v3.espacenet.com/textdraw?DB=EPODOC&IDX=WO2006082390&F=0&QPN=WO2006082390 ]


Original Document of WO2006082390
---------------------------------

[See European Patent Office Document
     http://v3.espacenet.com/origdoc?DB=EPODOC&IDX=WO2006082390&F=0&QPN=WO2006082390 ]


INPADOC legal status of WO2006082390
------------------------------------

No legal data found.

--------
(c) 2006 European Patent Office

[Return to top]

------------------------------

Date:    Wed, 13 Sep 2006 12:13:08 -0400
From:    "Bernice A. Melsky" <bernicemelsky@xxxxx.xxx>
Subject: RES: Iraqi Gulf War Veteran Refugees in the U.S.:PTSD and  Physical Symptoms

Iraqi Gulf War Veteran Refugees in the U.S.:PTSD and Physical Symptoms.

Soc Work Health Care. 2006;43(4):85-98.

Jamil H, Nassar-McMillan SC, Salman WA, Tahar M, Jamil LH.
15400 W McNichols, 2nd Floor, Detroit, MI, hjamil@med.xxxxx.xxx.

PMID: 16966311

Veterans of the Gulf War present various symptoms and maladies. Reports by
governmental and private entities have yielded mixed results and have been
fraught with criticisms of biased research design.

The vast majority of these studies have focused on U.S. veterans, with a
much smaller number focusing upon British veterans. Very few have examined
Iraqi Gulf War veterans.

Our study involves administering a health issues questionnaire to a sample
of Iraqi Gulf War veteran refugees in the U.S.

Results indicate relationships between Post-Traumatic Stress Disorder
(PTSD) scores and health outcome measures of chronic fatigue, fibromyalgia,
functional status, quality of life, and health care utilization in terms of
frequency and level of intensity.

Implications for further inquiry are presented.

[Return to top]

------------------------------

Date:    Fri, 15 Sep 2006 13:54:19 -0400
From:    "From Dr. Charles Shepherd <charlesbshepherd@xxxxx.xxx> via Co-Cure
         Moderator" <ray@xxxxx.xxx>
Subject: NOT,MED:  ME Association Medical Meeting in Chester:  Saturday September 9th


MAY BE REPOSTED

The ME Association held another very successful medical meeting in Chester
on Saturday September 9th.  Over 100 people reserved seats and with more
turning up than had booked we then had to bring in extra seating at the
last minute - so filling the hall to almost maximum capacity.

Unfortunately, Dr Anne Macintyre, who has special expertise in the area of
medical assessments for benefit purposes, was unwell and had to pass on her
apologies on Saturday morning for being unable to attend this year.  We
wish Anne a speedy recovery.

Dr William Weir, a specialist in infectious diseases from London, opened
the meeting by giving a presentation in which he set out the case for the
biomedical model of ME/CFS.  At the same time he carefully took apart the
psychosocial model involving abnormal illness beliefs and behaviour.

After a short interval Dr Charles Shepherd reviewed a number of national
'hot topics'.  These included

The MRC research agenda
The Gibson Inquiry
NICE Guidelines
Proposed new DWP medical guidance on ME/CFS for benefit purposes
New NHS services in England for people with ME/CFS.

Dr Weir and Dr Shepherd then answered a wide range of questions from the
audience.  Among the topics discussed were:

The role of CBT in the management of ME/CFS
The overlap with autoimmune and rheumatic conditions, including Sjogren's
syndrome
Controversies surrounding the diagnosis and management of Lyme disease
Progress towards a diagnostic test for ME/CFS
Ampligen
Immunotherapy, including interferon and immunoglobulin
Lightning Process
Osteopathic treatment
The pros and cons of vaccinations
Genetic factors which contribute towards ME/CFS
A more detailed account of the meeting will appear in the next (October)
issue of ME Essential.  This will also contain an in depth review of the
controversies surrounding Lyme disease and ME/CFS.  This has been written
by Dr Darrel Ho-Yen - a consultant microbiologist with a special interest
in both Lyme disease and ME/CFS.

An audio recording of the Chester meeting has also been made.  If demand is
there for copies we will consider making them available for a small
charge.  If you are interested in obtaining a copy please contact Tony
Britton at: anthonybritton536@btinternet.com.

The venue for the next MEA medical meeting has yet to be decided and MEA
trustees would like to hear from local groups in any part of the UK who
would be interested in being involved in one of these patient-orientated
half day meetings.


ENDS

[Return to top]

------------------------------

Date:    Sat, 16 Sep 2006 14:16:21 +0200
From:    "Dr. Marc-Alexander Fluks" <fluks@xxx.xx>
Subject: NOT,RES: European Patent Office has just changed the biography for the Gow et al. CFS patent

The European Patent Office has just changed the biography for the Gow et
al. CFS patent,
   http://v3.espacenet.com/textdoc?DB=EPODOC&IDX=WO2006082390&F=0
   https://listserv.surfnet.nl/scripts/WA.EXE?A2=ind0609B&L=me-net&P=R6243
   http://listserv.nodak.edu/cgi-bin/wa.exe?A2=ind0609B&L=co-cure&P=R3066

 * The layout is different.
 * There are more 'cited documents', there are links to (empty) documents.

Below you will find the updated 'Bibliographic data'...

---------------------------------------------------------------------------
Bibliographic data of WO2006082390
----------------------------------

Publication number: WO2006082390
Publication date:   2006-08-10
Inventor:           GOW JOHN (GB);
                    CHAUDHURI ABHIJIT (GB)
Applicant:          UNIV GLASGOW (GB);
                    GOW JOHN (GB);
                    CHAUDHURI ABHIJIT (GB)
Classification:
  - international:  C12Q1/68; G01N33/53; C12Q1/68; G01N33/53;
  - european:
Application number: WO2006GB00332 20060201
Priority number(s): GB20050002042 20050201

View INPADOC patent family

Cited documents:    XP002361324
                    XP002377235
                    XP002377236
                    XP005062766
                    XP002377237
                    XP002377238
                    XP002377239
---------------------------------------------------------------------------

Notes on the cited documents...

1. WO2006GB00332
   This is identical to WO2006082390.
2. GB20050002042
   This has not been listed.
3. XP002361324
   This doesn't seem to exist.
4. XP002377235
   This has not been listed.
5. XP002377236
   This has not been listed.
6. XP005062766
   This has not been listed.
7. XP002377237
   This has not been listed.
8. XP002377238
   This has not been listed.
9. XP002377239
   This has not been listed.

[Return to top]

------------------------------

Date:    Sat, 16 Sep 2006 15:22:01 -0400
From:    "Bernice A. Melsky" <bernicemelsky@VERIZON.NET>
Subject: RES: The health effects of at-home written emotional  disclosure in fibromyalgia: a randomized trial

The health effects of at-home written emotional disclosure in fibromyalgia:
a randomized trial.

Ann Behav Med. 2006 Oct;32(2):135-46.

Gillis ME, Lumley MA, Mosley-Williams A, Leisen JC, Roehrs T.

Wayne State University.

PMID: 16972811


Background: The presence and severity of the chronic pain syndrome
fibromyalgia (FM) is associated with unresolved stress and emotional
regulation difficulties. Written emotional disclosure is intended to reduce
stress and may improve health of people with FM.

Purpose: This study tests the effects of at-home, written emotional
disclosure about stressful experiences on the health of people with FM and
uses multiple follow-ups to track the time course of effects of disclosure.

Methods: Adults with FM (intention-to-treat, n = 83; completers, n = 72)
were randomized to write for 4 days at home about either stressful
experiences (disclosure group) or neutral time management (control group).
Group differences in immediate mood effects and changes in health from
baseline to 1-month and 3-month follow-ups were examined.

Results: Written disclosure led to an immediate increase in negative mood,
which did not attenuate across the 4 writing days. Repeated-measures
analyses from baseline to each follow-up point were conducted on both
intention-to-treat and completer samples, which showed similar outcomes. At
1 month, disclosure led to few health benefits, but control writing led to
less negative affect and more perceived support than did disclosure. At
3-month follow-up, these negative affect and social support effects
disappeared, and written disclosure led to a greater reduction in global
impact, poor sleep, health care utilization, and (marginally) physical
disability than did control writing. Interpretation of these apparent
benefits needs to be made cautiously, however, because the disclosure group
had somewhat poorer health than controls at baseline and the control group
showed some minor worsening over time.

Conclusions: Written emotional disclosure can be conducted at home, and
there is tentative evidence that disclosure benefits the health of people
with FM. The benefits, however, may be delayed for several months after
writing and may be of limited clinical significance.

[Return to top]

------------------------------

Date:    Sat, 16 Sep 2006 15:25:07 -0400
From:    "Bernice A. Melsky" <bernicemelsky@xxxxx.xxx>
Subject: RES: Synthesis and activity of a new class of dual acting norepinephrine and serotonin reuptake inhibitors: 3-(1H-indol-1-yl)-3-arylpropan-1-amines

Synthesis and activity of a new class of dual acting norepinephrine and
serotonin reuptake inhibitors: 3-(1H-indol-1-yl)-3-arylpropan-1-amines.

Bioorg Med Chem. 2006 Sep 12; [Epub ahead of print]

Mahaney PE, Vu AT, McComas CC, Zhang P, Nogle LM, Watts WL, Sarkahian A,
Leventhal L, Sullivan NR, Uveges AJ, Trybulski EJ.

Chemical and Screening Sciences, Discovery Medicinal Chemistry, Wyeth
Research, 500 Arcola Road, Collegeville, PA 19426, USA.

PMID: 16973367


Compounds with a combination of norepinephrine and serotonin reuptake
inhibition have been approved in the US and Europe for a number of
indications, including major depressive disorder and pain disorders such as
diabetic neuropathy and fibromyalgia. Efforts to design selective
norepinephrine reuptake inhibitors based on SAR from the aryloxypropanamine
series of monoamine reuptake inhibitors have led to the identification of a
potent new class of dual acting norepinephrine and serotonin reuptake
inhibitors, namely the 3-(1H-indol-1-yl)-3-arylpropan-1-amines.

[Return to top]

------------------------------

Date:    Sun, 17 Sep 2006 11:34:41 +0200
From:    "Dr. Marc-Alexander Fluks" <fluks@xxx.xx>
Subject: RES,NOT: CFS Treatment - Clinical Trials at the University of Cincinnati

Souce: Clinical Trial Studies
Date:  September 16, 2006
URL:   http://clinicaltrials.gov/show/NCT00375973
       http://communitydispatch.com/artman/publish/article_6431.shtml


Chronic Fatigue Syndrome Treatment: A Double Blind Trial of Duloxetine
By Clinical Trials and University of Cincinnati
----------------------------------------------------------------------
This study is currently recruiting patients.
Verified by University of Cincinnati September 2006

Sponsors and Collaborators:    University of Cincinnati
                               Eli Lilly and Company
Information provided by:       University of Cincinnati
ClinicalTrials.gov Identifier: NCT00375973


Purpose

The purpose of this study is to determine the safety and efficacy of
duloxetine compared with placebo for reducing fatigue in patients
diagnosed with Chronic Fatigue Syndrome (CFS).

Condition:      Chronic Fatigue Syndrome (CFS)
Intervention:   Drug: Duloxetine
Phase:          Phase II
                Phase III
Study Type:     Interventional
Study Design:   Treatment, Randomized, Double-Blind, Placebo Control,
                Parallel Assignment, Safety/Efficacy Study

Official Title: A Randomized, Placebo-Controlled, Double-Blind Trial of
Duloxetine in the Treatment of Patients With Chronic Fatigue Syndrome.

Further study details as provided by University of Cincinnati:
Primary Outcomes: Multidimensional Fatigue Inventory (MFI)
Secondary Outcomes: Efficacy:; Brief Pain Inventory (BPI); Hospital
Anxiety and Depression Scale (HADS); Clinical Global Impression of
Severity (CGI-S); Patient Global Impression of Improvement (PGI-I);
Safety:; Discontinuation rates; treatment-emergent adverse events; vital
signs; laboratory analyses

Expected Total Enrollment: 60

Study start: September 2006;  Expected completion: September 2009

Chronic fatigue syndrome (CFS) is characterized by severe disabling
fatigue of at least six months duration that cannot be fully explained by
an identifiable medical condition . Pain symptoms are also a part of the
diagnostic criteria for CFS, and include muscle pain, multi-joint pain,
and headaches. The prevalence of CFS ranges from 0.007 to 2.8 % in the
general adult population and 0.006 to 3.0% in primary care practice (2).
Although most who receive a CFS diagnosis are 30-40 years of age,
Caucasian, and female, CFS affects both women and men, adults and
children, and all racial and socioeconomic classes.

Patients with CFS have 2-4 times the rate of depression and anxiety
compared with the general population. CFS is also commonly comorbid with
fibromyalgia, a disorder characterized by chronic widespread pain,
tenderness, fatigue, sleep and mood disturbances. In some samples, 70% of
patients with fibromyalgia also meet criteria for CFS. CFS and
fibromyalgia are characterized by greater similarities than differences
and may share pathophysiologic features. Like fibromyalgia, CFS is
associated with chronic pain, sleep and mood disturbances. Because
fibromyalgia responds to treatment with antidepressants, particularly the
dual serotonin and norepinephrine reuptake inhibitors, including
duloxetine, antidepressant trials in CFS are clearly needed.


Eligibility

Ages Eligible for Study: 18 Years-65 Years, Genders Eligible for Study: Both


Criteria

Inclusion Criteria:
 1. Female and male outpatients between 18-65 years of age.
 2. Meet criteria for revised CDC definition of CFS (at least 6 months
    of persistent fatigue that substantially reduces the person’s level
    of activity; 4 or more of the following symptoms that must occur with
    fatigue in a 6-month period: impaired memory or concentration, sore
    throat, tender glands, aching or stiff muscles, multijoint pain, new
    headaches, unrefreshing sleep, and post-exertional fatigue. Medical
    conditions that may explain the fatigue and psychiatric disorders,
    including eating disorders, psychotic disorders, bipolar disorder,
    melancholic depression, and substance abuse within 2 years of the
    onset of fatigue, are excluded).
 3. Provision of written informed consent for participation in the trial.
 4. Educational level and degree of understanding such that the patient
    can communicate intelligibly with the investigator and study staff.
 5. Judged to be reliable and agree to keep all appointments for clinic
    visits, tests, and procedures required by the protocol.

Exclusion Criteria:
 1. Current melancholic major depressive disorder, or a previous diagnosis
    of psychosis, eating disorder, or bipolar disorder.
 2. History of substance abuse or dependence within the past year, excluding
    nicotine and caffeine.
 3. A positive urine drug screen for any substance of abuse (may be retested
    if positive test was for a prescribed medication that was not washed out).
 4. Women who are pregnant or breast feeding; women must test negative for
    pregnancy at Visit 1.
 5. Women of childbearing potential who are not using a medically accepted
    means of contraceptive when engaging in sexual intercourse.
 6. Patients who, in the opinion of the investigator, are treatment-refractory
    or whose response is likely to be compromised by existing or future
    disability compensation issues.
 7. Serious unstable medical illness, including cardiovascular, hepatic,
    renal, respiratory, or hematologic illness, or other unstable medical
    or psychiatric conditions that in the opinion of the investigator would
    compromise participation or would likely lead to hospitalization during
    the duration of the study. Abnormal TSH concentrations (unless treatment
    for hypothyroidism has been stable for at least the past 3 months and the
    patient is clinically euthyroid).
 8. Patients who have uncontrolled narrow-angle glaucoma.
 9. Patients who have acute liver injury (such as hepatitis) or severe
    cirrhosis (Child-Pugh Class C).
10. Patients who are judged prior to randomization to be at suicidal risk by
    the clinical investigator.
11. Treatment with antidepressant medication within 14 days prior to
    randomization with the exception of fluoxetine, which cannot be used
    within 30 days prior to randomization. Potential need to use a MAOI
    during the study or within 2 weeks of discontinuation of study treatment.
12. Patients who have previously taken duloxetine
13. Patients who are taking any excluded medications that cannot be
    discontinued at Visit 1.
14. Treatment within the last 30 days with a drug that has not received
    regulatory approval at the time of study entry.
15. Known hypersensitivity to duloxetine or any of the inactive ingredients.


Location and Contact Information

Please refer to this study by ClinicalTrials.gov identifier  NCT00375973
Jennifer M Hoff, MSW, LSW   513-475-8114  jennifer.hoff@xx.xxx
Susie G Sheridan, B.S.      513-475-8115  dana.sheridan@xx.xxx

Ohio
Psychiatric Professional Services Inc., Women's Health Research
Program, Cincinnati,  Ohio,  45219,  United States; Recruiting
Jennifer M Hoff, MSW, LSW   513-475-8114  jennifer.hoff@xx.xxx
Susie G. Sheridan, B.S.     513-475-8115  dana.sheridan@xx.xxx
Lesley M. Arnold, MD, Principal Investigator


Study chairs or principal investigators

Lesley M Arnold, MD,  Principal Investigator,  University of Cincinnati
Women's Health Research Program

--------
(c) 2006 Clinical Trials

[Return to top]

------------------------------

Date:    Sun, 17 Sep 2006 11:33:38 +0200
From:    "Dr. Marc-Alexander Fluks" <fluks@xxx.xx>
Subject: RES,NOT: FDA Asks U.S. Marshals to Seize FM Dietary Supplements

Source: EmaxHealth
Date:   September 14, 2006
URL:    http://www.emaxhealth.com/83/7383.html


FDA Asks U.S. Marshals to Seize Dietary Supplements
---------------------------------------------------

Dietary Supplements Fals Claim

At the request of the U.S. Food and Drug Administration (FDA), on
September 5, 2006, U.S. Marshals seized quantities of Ellagimax capsules,
Coral Max capsules, Coral Max without Iron capsules, and Advanced
Arthritis Support capsules distributed by Advantage Nutraceuticals L.L.C.
of Fort Walton Beach, Florida, valued at approximately $55,000.00.

The products are alleged to be in violation of the new drug and
misbranding provisions of the Federal Food, Drug and Cosmetic Act.

Although these products are labeled as "dietary supplements," they are
being promoted with claims typically associated with drug products. These
claims are evident in the products' labeling, including various
promotional literature, an audio cassette tape, and two Internet Web sites
owned and used by the firm. We advise consumers to consult with their
physician if they are taking these products.

Advantage Nutraceuticals' labeling promotes and markets these products for
serious disease conditions, including but not limited to cancer,
arthritis, fibromyalgia and seizures.

FDA considers these products to be unapproved new drugs, and the agency
takes seriously its responsibility to protect Americans from dangerous
unapproved drugs. Before a new drug product may be legally marketed, it
must be shown to be safe and effective and approved by FDA. Drug product
labeling must also include adequate directions for use, which the seized
products' labeling does not provide.

Following an investigation of the firm's marketing practices, FDA advised
Advantage Nutraceuticals that the claims related to prevention or
treatment of diseases in many of its products' labeling make these
products subject to regulation as drugs. Despite FDA's warnings, the firm
failed to take sufficient steps to come into compliance with the Act.
During subsequent inspections, FDA inspectors found that the offending
claims were still being made.

--------
(c) 2006 EmaxHealth

[Return to top]

------------------------------

Date:    Sun, 17 Sep 2006 11:29:04 +0200
From:    "Dr. Marc-Alexander Fluks" <fluks@DDS.NL>
Subject: RES,NOT,URL: CFS Belief Systems

Today, an ME-NET web page has been created, entitled,
   CFS Belief Systems
The URL is,
   http://www.me-net.dds.nl/meweb/web4.12.html

The present page is just the first version. Please send your input and
feedback to <me-net@dds.nl> for future updates.

[Return to top]

------------------------------

Date:    Mon, 18 Sep 2006 16:10:37 -0400
From:    Ken Coleman via Co-Cure Moderator <ray@xxxxx.xxx>
Subject: Re: RES,NOT: CFS Treatment - Clinical Trials at the University of Cincinnati

Upon searching for DULOXETINE; I found the following information at
<http://www.medscape.com/druginfo/search?cid=med&queryText=Duloxetine&submit=Search&search_type=drug>

I Seriously recommend anyone interested in this announced study to
first consult the information on Medscape at the above URL!


 From the URL above:
         Industry Information

         CYMBALTA® (duloxetine HCI) Infosite - Broad and rapid
         relief of both emotional and physical symptoms of
         depression in as early as 1 to 2 weeks of treatment.
         CYMBALTA helps patients achieve high rates of remission.
         For Prescribing Information, Boxed Warning and Safety
         Information, and important findings, click here.

         CYMBALTA® (duloxetine HCl) InfoSite - The 1st FDA-approved
         treatment for the management of diabetic peripheral
         neuropathic pain. CYMBALTA provides effective pain relief
         with once-daily dosing. Learn more about CYMBALTA.

                   Review Boxed Warning & Safety Information and
                   Prescribing Information.

         Drugs matching Duloxetine:
                 Duloxetine Oral (generic)

         Recalls and Warnings related to Duloxetine

         Medscape Alert -
         Cymbalta May Aggravate Preexisting Liver Disease10/18/2005
         Medscape Alert -
         FDA Warns Against Use of Migraine Drugs With Certain Anti
         depressants7/20/2006
         Medscape Alert - FDA Issues Public Health Advisory for Antidepressant
         Use in Children and Adolescents10/15/2004


Ken Coleman
Email: m.e.s.s@xxx.xxx


[Return to top]


End of CO-CURE Medical & Research Posts Only Digest - 11 Sep 2006 to 18 Sep 2006 (#2006-43)

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