Part One - Unmasked Research:
STAT1-alpha and p53 Deficiencies are Found in Patients with
Chronic Fatigue Syndrome;
The National CFIDS Foundation Responds by Funding Several New Research
Investigative Report by Alan Cocchetto, Medical Director, NCF, Inc.
© 2003 Written Permission Required for Reprinting or Distributing
Through its own extensive six month medical investigation, the National CFIDS
Foundation (NCF) uncovered research that will have widespread ramifications
among the CFS patient community, their physicians, as well as CFS researchers
worldwide establishing CFS as a serious and perhaps fatal illness. Because of
these implications, the NCF has responded to this major discovery by dynamically
funding new medical research by several noted scientists.
Briefly stated, the NCF has uncovered a key scientific discovery, previously
made by prominent CFS researchers, that has yet to see the 'light of day.' The
critical component: STAT1. The STAT1 protein is crucial for proper immune
function and regulation. Without it, cells are unresponsive to interferons
leaving the body defenseless against viral and bacterial infections. In fact,
human STAT1 deficiency is lethal. In this report, the NCF identified CFS
researchers that previously found STAT1 to be absent in CFS patient blood.
"The NCF began its investigation by thoroughly reviewing work that had been
previously published by Kenny DeMeirleir, MD, PhD and his colleague's group in
Belgium" stated Gail Kansky, President of the NCF. "Then one discovery lead to
another and our own investigation just skyrocketed" said Kansky. "As the CFS
patient community is aware, Dr. DeMeirleir is a highly visible CFS researcher,
both in the U.S. and abroad. He is a medical editor for the Journal of Chronic
Fatigue Syndrome, is on the board of the American Association for Chronic
Fatigue Syndrome (AACFS), and is actively involved with R.E.D. Laboratories
which has worked to further the studies of Dr. Robert Suhadolnik, from Temple
University, whose research group made the initial RNase L discoveries in CFS
patients" stated Kansky. "I strongly feel that the NCF's investigational and
medical research efforts will greatly add to those efforts that we have
previously undertaken to unravel the mysteries of CFS. This should lead to much
greater understanding of the disease process" said Kansky. Furthermore, she
stated that "Our funded medical research is aimed to solidify and scientifically
expand this important work so that desperately needed and appropriate patient
treatments will follow. The NCF's efforts will continue to illuminate the
darkness that has overshadowed this very serious illness."
The NCF first began its investigation with a published book titled Chronic
Fatigue Syndrome: A Biological Approach edited by Dr. Patrick Englebienne and
Dr. Kenny DeMeirleir . Published in 2002, examination of the chapters, by NCF
staffers, revealed that Dr. DeMeirleir and his colleagues carefully and
methodically probed various signal transduction pathways in patients with CFS
via examination of their peripheral blood mononuclear cells (PBMC's or monocytes).
These authors stated that "the importance of STAT1 in mediating the action of
both type I and II IFNs (interferons) is no longer questioned, as a lack of its
expression is consistently associated with IFN resistance. The apparent
dysregulation in types I and II IFN pathways in chronic fatigue syndrome led us
to investigate the expression of STAT1 in PBMCs. We classified the samples
according to the ratio of 37- over 80- kDa RNase L which is representative of
the proteolytic activity of the PBMC samples. As shown in Figure 5.9, STAT1 is
fully degraded in positive samples, suggesting that it may also be a substrate
of the proteases responsible for RNase L cleavage. A degradation of STAT1 in
those cells might well constitute the missing link explaining unresponsiveness
to IFNs" (interferons) .
In this figure from the book, with RNase L ratios (37 kDa/80 kDa) greater than
0.25, native STAT1-alpha is lost but some cleavage (breakdown) products are
shown until the RNase L ratio hits approximately 2.0. After that, there is the
complete absence of both STAT1-alpha and the cleavage product! Explaining
further, STAT is notation for Signal Transducers and Activators of
Transcription. STAT's are a family of transcription factors that play central
roles in the responses of cells to cytokines, molecules that control every
aspect of the immune system . STAT1 has two forms, alpha and beta. Dr.
DeMeirleir's group tested STAT1-alpha, a native 91 kDa protein, in patients with
CFS and correlated it with the RNase L ratios. STAT1-alpha is intimately
involved in the response of cells to type I (alpha and beta) and type II (gamma)
interferons. Most important is the fact that a STAT1-alpha deficiency is
associated with fatal infections by both viruses and bacteria [3,4,5,6,7,8].
Furthermore, in animals, STAT1-alpha deficiency is associated with impaired
responses to interferons, increased susceptibility to tumors, as well as
impaired growth control. STAT1-alpha also plays a critically important role in
The NCF staff then wondered if Dr. DeMeirleir and his colleagues realized the
importance, as well as the significance, of their discovery and when might they
have first identified it? To answer this question, we uncovered a patent filed
in April 1999 titled "Methods and compositions for use in characterizing
multiple sclerosis disease activity in a subject."  In this patent, Dr.
DeMeirleir and his colleagues at R.E.D. Laboratories determined the amount of 37
kDa and 80 kDa RNase L proteins and utilized RNase L ratios to characterize
multiple sclerosis disease activity. This patent was filed after one filed by
inventor Dr. Robert Suhadolnik, in April 1999, titled "Chronic fatigue syndrome
diagnosis" in which the diagnosis of CFS was made through the detection and
determination of both the 37 kDa and 80 kDa RNase L proteins . Examination
of both these patents indicated that, in both CFS and MS, these same RNase L
proteins can be used to assist in diagnosis as well as for determining disease
activity. Multiple sclerosis patients have previously been found to have
alterations in STAT1-alpha . Furthermore, in CFS, this is vitally important
due to the correlations that were found between the RNase L ratios and
This is further stated in two additional patent applications by Dr. DeMeirleir's
colleagues. One of these applications directly discusses the specific role of
STAT1-alpha in CFS patients [12,13]. Quoting these patent applications directly:
"STAT1 plays an important role in growth arrest, in promoting apoptosis and is
implicated as a tumor suppressor. STAT1 null cells are resistant to apoptotic
induction by TNF-alpha....STAT1 deficient mice exhibit a severe defect in IFN-dependent
immune responses against viruses and microbial pathogens....If STAT1 is disabled
or otherwise inactive in the cells of the immune system, treatment with
interferon or interferon inducer will not be effective in promoting and
establishing the interferon-inducible antiviral and antiproliferative pathways."
Furthermore these applications state "the results demonstrate that the presence
and amount of STAT1 protein fragmentation directly correlates with the presence
and amount of low molecular weight RNase L fragments in PBMC samples. These data
indicate that native STAT1 protein is fragmented at an earlier point in the
disease cycle than RNase L, and that by the time native RNase L is demonstrably
attacked by proteases (ratio > 2.0), that native STAT1 protein has entirely
disappeared due to proteolysis, leaving the cells unable to respond to
interferons and/or interferon inducers."
It became obvious to NCF staffers that Dr. DeMeirleir and his colleagues had
made a vitally important discovery but had only published part of the
information in the book and the remainder in patents or patent applications that
we were aware of. Was there other information to be found?
A quick check of the R.E.D. Laboratories website  indicated that "The
company's scientists have discovered a number of other cellular proteins that
play important roles in the initiation, progression, and pathogenesis of immune
dysfunction. The most important revelation to date has been the discovery of one
protein, which is used as a serum-based assay for the detection of chronic
immune disease. Such a serum-based marker has been developed as a screening test
for the blood supply to reduce the number of transfusion-related infections with
persistent (and covert) viral infections."
It became obvious to the NCF staffers, that both the book and the patent
information told a significant story regarding the critical importance of
STAT1-alpha in CFS patient blood. One thing is certain, blood doesn't lie!
Likewise, the true implications, pathological and perhaps life threatening for
patients as well as financial potentially for R.E.D. Laboratories, regarding
transfusion-related infections in the blood supply by "covert infections", were
enormous and certainly worthy of worldwide attention!
Probing further, the NCF discovered vitally important testimony given directly
by Dr. Kenny DeMeirleir before the Flemish Parliament of Belgium in a hearing on
March 5, 2001 . In his testimony given before Flemish Parliament, Dr.
DeMeirleir stated "Caspases and calpain are induced by cellular stress, which
leads to apoptosis. In an intracellular disorder, calcium influx is increased.
Calcium will further activate calpain, so that some caspases are inhibited and
therefore block apoptosis. One of the cellular proteins that are split by these
enzymes is STAT1, the carrier of the interferon-gamma signal in the immune
cells, which leads to the Th1-to-Th2 shift. Unfortunately, the Chronic Fatigue
Syndrome is often called psychosomatic. This is, however, more indicative of
medicine's inability to deal with it. We now understand the nature of the
disorder. In its early phase, apoptosis increases. In a subsequent evolution,
apoptosis is blocked and the interferon signal disappears due to the destruction
of the protein which transports the intracellular interferon signal to the
nucleus (where apoptosis is initiated). This leads to more and more infections.
This process goes on at all levels (in the central nervous system, the muscle
cells, the white blood cells, and so on). Some patients suffering from the
Chronic Fatigue Syndrome develop epilepsy. We find that most patients have a
light form of epilepsy. This leads to sleep disturbances and a situation where
the fatigue increases since one does not recover anymore."
"Given the true significance of this work and its implication in a functionally
oppressive disease that desperately struggles for proper recognition,
validation, and affirmation at all levels of medical science, the NCF's Board of
Directors unanimously voted to immediately fund several important research grant
projects to be carried out by noted researchers" said Kansky.
The NCF first contacted a well-known research group, who has asked to remain
anonymous at this time, and arranged to send grant funding to them via our NCF
Research Grant Program. "We are very excited by this since this group will be
the first to begin studying "STAT1-alpha in Chronic Fatigue Syndrome Patients"
stated Kansky. Lead scientists for this research group commented that this was
an enormous step forward towards understanding the disease process in these
Next, the NCF arranged another research project, also funded by our Research
Grant Program. The NCF sent grant money to Dr. Konstance Knox and Dr. Donald
Carrigan, both from the Institute for Viral Pathogenesis, for "A Study for the
Potential Role of STAT1 in the Pathogenesis of Chronic Fatigue Syndrome." "This
grant is aimed at significantly expanding our knowledge about the specifics of
STAT1 and its role in CFS from a pathological standpoint" stated Kansky. "Futhermore,
since we are expecting both teams (first group and Knox/Carrigan) to complete
their work with research physicians in the months ahead, we are anticipating
significant medical breakthroughs as the direct result of these grants. The NCF
has expedited this work because our patients are seriously ill and we anticipate
that these efforts will certainly confirm this" said Kansky.
The NCF staff also contacted two world renown experts on the STAT1 protein: Dr.
Joan Durbin from Columbus Children's Research Institute and Dr. David Levy from
New York University School of Medicine. Both Dr. Durbin and Dr. Levy have
published extensively on STAT1 and are responsible for the development of the
animal model for STAT1-alpha deficiencies. In NCF interviews with both of them
and from their numerous published medical journal articles, it is clear that the
complete loss of the native STAT1 protein constitutes a serious illness that may
ultimately be fatal unless this protein defect can be corrected.
Ironically, this is where the NCF is truly hopeful. In one of the patent
applications , Dr. DeMeirleir's colleagues determined that the "STAT1
protein is degraded when a cell extract from a healthy control (i.e., 'negative
extract'; RNase L ratio < 0.2, STAT1 protein containing) is incubated with a
cell extract from a patient (RNase L ratio = 3.0; STAT1 protein negative). This
degradation is inhibited in the presence of proteasomal inhibitor (MG132) but
not in the presence of the other protease inhibitors tested. Thus the
degradation of STAT1 protein is a specific cellular process that involves
proteasome and does not involve the apoptotic enzymes caspase-3 or calpain."
What this implies is that Dr. DeMeirleir's colleagues had already found a very
specific inhibitor, MG132, for STAT1-alpha degradation in CFS patient blood
in-vitro! In our own research, the NCF found that MG132 functions as both a
specific inhibitor of the ubiquitin-proteasome pathway  as well as a
specific beta-secretase inhibitor . Both of these pathways are intimately
involved with amyloid formation. Amyloidogenesis is a general phenomenon of
protein misfolding. Protein misfolding and the formation of abnormal protein
fibrils appear to play a central role in a variety of neurodegenerative diseases
including Alzheimer's disease, Parkinson's disease, Huntington's disease, etc.
This is why the expression of the Huntington's protein in CFS patients, by the
CDC using gene expression analysis, is intriguing . These are all scientific
clues to a bigger picture for this disease.
Because of our previous efforts with the ciguatera epitope and its effects on
neuroblastoma cells as well as other specific unannounced scientific findings
related to this epitope, the NCF has also provided new research grant funding to
Dr. Yoshitsugi Hokama, from the University of Hawaii, for the "Development of an
Immunological Assay for Assessment of Amyloids in Chronic Fatigue Syndrome."
"Dr. Hokama's previous research dovetailed nicely into the scientific work we
are describing here" said Kansky. "In fact, the NCF believes there is a
relationship between the ciguatera epitope, protein misfolding, and amyloid
Amyloids directly alter sodium channels and that is part of the significance of
the ciguatera epitope finding as it relates to our previous CFS patient studies.
This is something that we had known prior to the initial funding of Dr. Hokama's
work but this previous work had to be scientifically verified and expanded upon
first before we pressed on further" stated Kansky. The NCF is also aware that a
relationship already exists between amyloid formation and STAT1-alpha. Amyloids
can potentially bind the STAT1-alpha protein and take it out of commission. This
mechanism is very important. Dr. DeMeirleir and his colleague's discovery on
STAT1-alpha and RNase L cleavage is central to our working hypothesis but the
NCF believes that we have found other missing pieces, regarding the infectious
agent as well as other disease mechanisms, that are absent from their work. What
is exciting is that our NCF staff has already identified very specific PCR
binding sequences for STAT1-alpha in the infectious agent that we believe is
directly responsible for this disease. This implies that once infection begins,
STAT1-alpha becomes one of the first critical immune system proteins to get
"chewed up" as a result of being a direct target of the infectious agent thereby
greatly damaging the immune system! This would be in agreement with Dr. De
Meirleir's findings where STAT1-alpha binding occurs before the degradation of
the native RNase L protein.
"Furthermore, in full agreement with Dr. Elaine DeFreitas' previous work, the
NCF believes this infectious agent is of non-human origin" stated Kansky. "We
are meeting the dragon head-on and we hope that our funded research will help
crack the code, once and for all, for this disease. The NCF continues to break
important new ground in the understanding of the disease mechanisms while going
scientifically where no other CFS organization has ever been before! This
appears to be our forte and it reflects our true passion and full committment to
the CFS patient community. As a result, we are much further ahead in the
discovery process than most patients and researchers realize" said Kansky.
In addition, Dr. DeMeirleir's colleagues also made another significant discovery
critical to the health and welfare of patients with CFS [19,20]. Excerpts from
the book and the patent application state "Fig. 1 represents a densitometric
scan of a Western blot detecting p53 protein and p53 protein fragments from PBMC
extracts from CFS patients....The above results demonstrate that the presence
and amount of p53 protein fragmentation directly correlates with the presence
and amount of low molecular weight RNase L fragments in PBMC samples. These data
indicate that native p53 protein is fragmented at a later point in the disease
cycle than RNase L protein. The loss of functional p53 protein in PBMCs render
these cells unable to respond to normal growth inhibitory stimuli and provide
the means whereby unregulated cell growth occurs, ultimately giving rise to
Furthermore, their explanation in this application states: "Another important
protein that regulates the induction of apoptosis is p53. The p53 protein is
normally activated in response to genetic damage within the cell and its
activation is accompanied by self-stabilization, allowing it to accumulate to
high levels and cause cell cycle arrest and induce apoptosis. In addition, the
p53 protein has a critical role in protecting the cell from malignant
development; mutations in the p53 gene (and protein) are the most frequently
detected genetic event in cancer.
Mutations in p53 may occur at the genetic level (i.e. DNA sequence alterations
that change the amino acid structure of the protein), or its function may be
altered by alterations in the numerous proteins with which p53 interacts. p53
may also be altered by the action of certain proteases, resulting in cleavage,
preventing the formation of active tetramers of the protein. If p53 is cleaved
and/or otherwise disabled in the cells of the immune system, these cells would
be blocked from entering the apoptotic pathway if infected with a virus or other
microorganism. In addition, persistent inactivation of the p53 protein may lead
to increased incidence of cancer (Levine et. al., J. Chronic Fatigue Syndrome 7:
29-38, 2000). Activation of the 2-5A synthetase / RNase L antiviral pathway has
been demonstrated to induce apoptosis, while induction of the same pathway in
cells expressing mutant forms of p53 was demonstrated to suppress the apoptotic
pathway. The inactivation of p53, RNase L, and other proteins within the cells
of the immune system most certainly leads to a dysfunctional immune system,
unable to respond to challenge by microorganisms and/or the presence of
Indeed the immune system itself may be in a pre-malignant state."
In a review of the literature on STAT1-alpha and p53, we found an article 
on STAT1 deficient mice that states "when the STAT1 deficiency was placed on a
p53 null background there were increased rates of tumor formation and an
increase in the non-lymphoid tumor types." This helps to explain the importance
of these two proteins that are essential to appropriate immune responses since
STAT1 and interferon are central to antitumor effects . In fact, this study
suggests that NK (Natural Killer) cells from STAT1 deficient mice may have a
reduced capacity to eliminate tumor cells in-vivo.
Part one article review at a glance:
Dr. DeMeirleir and colleagues scientific finding:
STAT1-alpha deficiency in CFS patients
p53 deficiency in CFS patients
Loss of STAT1-alpha constitutes a serious illness that may ultimately be fatal
because the cells are unresponsive to interferon leaving them unable to
adequately defend against infections.
Loss of p53 constitutes a pre-malignant state because surveillance against DNA
mutations, protein alterations, and unregulated cell growth/division are left
Loss of these proteins assists in immune deficiency and dysregulation.
[Ed. Note: The references for this article have been combined with the
article, "Total Exposure."]