Ongoing ME or CFS related XMRV studies

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Archive of Ongoing ME or CFS related XMRV studies


UK/ University College London

  • Dept/School: Division of Infection & Immunity, University College London
  • Project Supervisor(s): Prof G Towers & Prof P Kellam
  • Laboratory supervisor: Prof Greg Towers
  • Clinical supervisor: Prof Deenan Pillay


Xenotropic murine retrovirus (XMRV) has recently been associated with chronic fatigue syndrome as well as prostate carcinoma in humans (1-3). XMRV is a murine endogenous virus found in the genome of mice and until recently has been thought to be absent from the human population. It is now becoming clear that XMRV has transmitted to humans by a process of zoonosis, presumably from mice, and appears to be associated with a variety of diseases not previously associated with viral infection.


1. We will establish quantitative PCR assays and serology assays including enzyme linked immunosorbant assays (ELISA) to detect and quantify XMRV. Importantly, assays used to detect related murine leukaemia viruses in the lab are expected to be suitable.

2. We will use these assays to measure XMRV load in chronic fatigue patient samples as well as, well but XMRV infected control samples, with a view to establishing whether viral load relates to disease, episodes of illness and/or severity.

3. The receptor for XMRV has been identified. We will seek human polymorphism in the xenotropic receptor and assess which human cells express it. We will also establish which cells in vivo in blood express the receptor and which cells are infected with XMRV by quantitative PCR on sorted subsets of B and T cells from XMRV infected individuals.


This project proposes to address some of the most important questions surrounding the recently described XMRV infection of humans and to seek a therapeutic strategy for XMRV treatment. We expect it to be a competitive project and the experiments performed are likely to be influenced by ongoing studies published as we go. We expect that the candidate will be fully trained in modern techniques of molecular virology during the course of this project.



USA/ Alter, NIH: 1000 blood donors study

  • Harvey J. Alter, MD, chief of clinical studies and associate director for research in the Department of Transfusion Medicine at the NIH Clinical Center in Bethesda, Md.

Looking for virus and antibodies in samples from 1000 donors. They also have frozen samples taken from blood recipients before and after transfusion and have linked them to donors. They will be looking to see if they can demonstrate any transmission.


This study may be being conducted in the Red Cross. (Mentioned on CAA webinar of the 17 December on Phase II results of the Blood Working Group] [1]



USA/ Hanson, Cornell University Ithaca/NIAID funding

  • Contact Principal Investigator: HANSON, MAUREEN REBECCA
  • Awardee Organization: CORNELL UNIVERSITY ITHACA
  • Administering Institutes or Centers: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
  • Total Funding: $269,496


  • Study Section: Special Emphasis Panel (ZRG1)
  • Fiscal Year: 2010
  • Award Notice Date: 8-JUN-2010
  • Project Start Date: 15-JUN-2010 Project End Date: 31-MAY-2012
  • Budget Start Date: 15-JUN-2010 Budget End Date: 31-MAY-2011


Abstract:

DESCRIPTION (provided by applicant):

Chronic fatigue syndrome (CFS) is an illness characterized by long-term fatigue, impaired memory or concentration, sore throat, tender lymph nodes, muscle pain, multi-joint pain, new headaches, unrefreshing sleep, and exercise intolerance. While the cause(s) of chronic fatigue syndrome has not been established definitively, a number of outbreaks implicate an infectious etiology. A variety of pathogens have been reported in individual CFS patients or found more frequently in CFS patients than controls, including various types of viruses. The immune systems of CFS patients exhibit both chronic activation and dysfunction. Recently, a new human retrovirus named XMRV (Xenotropic Murine Leukemia Virus-Related Virus) has been detected in a large proportion of CFS patients tested, but in only 4% of healthy subjects. We plan to learn more about the association of XMRV and other pathogens with CFS by examining an outbreak cohort not previously screened for the presence of XMRV. Other viruses, microbes, and parasites that may be associated with XMRV will also be detected with the use of a panmicrobial DNA microarray. We will determine whether the presence of XMRV and/or other pathogens is related to the current state of health of individuals who became ill during a 1985 outbreak in rural New York. Assuming that most members of the New York cohort are infected with XMRV, we will amplify and sequence XMRV envelope genes derived from blood cells of 40 individuals who became ill as children in 1985. We will also sequence envelope genes from 80 subjects in the Nevada cohort known to exhibit a high degree of XMRV infection. We will examine phylogenetic relationships between the Nevada and New York XMRV variants and will observe whether any amino acid substitutions correlate with the current state of health of the subjects. We will also study the possible association of XMRV and XMRV protein expression in the phenomenon of exercise intolerance. We will examine XMRV-infected CFS patients before and after an exacerbation of symptoms caused by serial exercise testing. We will determine whether increases in inflammatory cytokines, a growth factor, and nitric oxide, or blood cell changes are correlated with the extent of reduced physical ability that can be measured during a second exercise test taken after induction of postexertional malaise. All of these experiments will be designed to assess whether particular XMRV sequences and expression levels play a role in the symptoms experienced by CFS patients and to detect possible underlying mechanisms of the disability that impairs their physical activity.


PUBLIC HEALTH RELEVANCE:

Chronic fatigue syndrome, an illness that disables many Americans, has recently been associated with the presence of a new human retrovirus. We will investigate whether this retrovirus is both necessary and sufficient for the development of the illness, or whether additional pathogens are involved. We will determine whether the level of health and exercise intolerance in chronic fatigue syndrome is related to particular virus variants, expression of viral proteins, and/or dysfunction of the immune system.



USA/ Blood XMRV Scientific Research Working Group

Four phase study to assess the risk to the blood supply. Six labs are participating—the CDC, FDA (2), National Cancer Institute (NCI), WPI, and Blood Systems Research Institute (major contractor for the REDS 2 study).


Phase 1 results already published, Phase II results to be reported at the start of November 2010.

Phase 2 results announced on 14 December 2010, published by CAA on the 17 December 2010, Phase III results to be reported early 2011. [2]


Please go to the Blood XMRV Scientific Research Working Group page for further details.


USA Government Study/Lipkin

NIH director Francis Collins, has asked Anthony Fauci, head of the National Institute of Allergy and Infectious Diseases, to direct a multi-center study of CFS patients to get to the bottom of the dispute. Fauci says he asked W. Ian Lipkin, a professor of epidemiology at Columbia University’s Mailman School of Public Health, to head up the study.


The new study will involve fresh blood samples from 100 CFS patients and 100 similar, but healthy people — 25 of each group from four different sites around the country, to provide geographic diversity. The samples will be processed, blinded and sent to the FDA, the CDC and the Whittemore Peterson Institute. If a lab finds a sample is positive for XMRV, further tests will be needed to confirm the result. If one lab finds a positive sample but another lab doesn’t, the same samples can be shipped again, with a new blinded code, to be tested again. If the they get the same result, it is valid.


At least three labs have agreed to test fresh blood samples. The WPI, FDA/NIH and CDC. (The CDC is the only one that has not found MLV-related retrovirus)


Patients and healthy controls will come from clinicians in Miami, Boston, Palo Alto, and Salt Lake City. And they will be selected using the Fukuda and Canadian definitions, as well as showing signs of infection around the time they developed CFS. (Such as a sore throat or tender lymph nodes) [3]



UK/ Imperial College London (McClure)/HPA/NHSBT

Not sure if this involves ME or CFS.


The post is full-time and is fully funded, without clinical commitments, by the National Health Service Blood and Transplant Board. NHSBT has a need to generate data on XMRV relevant to transfusion practice in England and seeks to appoint a clinical trainee with an interest in microbiology or virology to provide clinical insight and support of collaborative work ongoing between the NHSBT, HPA and Imperial College London.


The successful candidate will be expected to assist with ongoing projects involving generating a better understanding of the implications of XMRV and the related PMRV agents for transfusion and transplantation practices in England. It is expected that the Clinical Research Fellow will apply clinical experience to develop research interests utilising the contacts afforded by the existing collaborations.



USA Retrospective and Prospective Donor MRV Surveillance and Transmission Studies

This study was mentioned at the Blood Products Advisory Committee Meeting on the 14 December 2010.[4]


Retrospective Study

5 repositories involved:

  • Transfusion Transmitted Viruses Study (TTVS) - Donor/Recip, Serum.
  • Transfusion Safety Study (TSS) - Donations, Serum.
  • Retrovirus Epidemiology in Donors Study (REDS) General Leukocyte and Plasma Repository (GLPR) - Donations, Plasma; Frozen Whole Blood
  • Viral Activation by Transfusion Study (VATS) - Donor/Recip, Plasma; Frozen Whole Blood
  • REDS Allogeneic Donor and Recipient Repository (RADAR) - Donor/Recip, Plasma; Frozen Whole Blood


AIMS:

  • It will use a total of 10,000 samples from 5 NHLBI repositories (2000 from each), collected over 4 decades.
  • They wil look at rates of transmission and correlates of transmission (is transmission associated with donations that have nucleic acid/antibody positivity)
  • Look at transfusion/transmission rates.
  • Effect of routine blood filtration (leukoreduction) and blood component storage period on transmission
  • Look at storage time of blood on infectivity
  • Looking for data on mortality and morbidity (It will be limited)
  • Should allow them to determine if virus has been expanding in the donor population over those four decades, or has it been around at a stable level for decades.
  • Will look at age, gender, ethnicity, geographic regions, etc.
  • Will look at acquisition risk (underlying diseases, surgery, etc)
  • Will look at course of infection and the clinical outcome of recipients if identified
  • Compare virus in donor to recipient.
  • Look at levels of virus, diversity of virus, seroconversion, etc.
  • Should help guide insights into the pathogenesis of the infection in general.
  • Utilize Abbott and Gen-Probe high-throughput screening assays for serology and NAT. (Need those who can perform large throughput of samples)
  • Assuming prevalence rates of 0.2% to 7% (Based on positive studies), we would identify 20 to 700 blood donors with viremia (NAT +) and/or evidence of XMRV/MLV exposure (antibody +), which will allow us to establish temporal and demographic trends of infection in the US donor population over the past 4 decades.


Prospective Study

It is too early to begin, and they don't have the tests yet. But have already begun to plan for this and think about funding:

  • Prospective screening of 20,000+ blood donors for XMRV/MLVs
  • Parallel NAT (e.g., Gen-Probe/Novartis TIGRIS) and antibodies (e.g., Abbott Architect) assays performed prospectively in donor screening labs under FDA Investigational New Drug (IND) exemption
  • Interdiction of XMRV/MLV-reactive blood components with deferral of donors
  • Enroll donors into follow-up study with immunologic and virologic assessments (in parallel with CFS patients)
  • Lookback investigations to determine XMRV status of recipients of previous units from XMRV+ repeat donors
  • What are the criteria to justify a prospective trial?


USA/ American Red Cross (ARC)/Cross section and repository studies/Stramer & Dodd

According to Michael Busch, on the CAA webinar which covered the Phase II results of the Blood Working Group (17 December 2010), American Red Cross (ARC) employees Roger Dodd Susan Stramer, are conducting cross section and repository studies.[5] No further details are known.


German/Dr Bieger

Testing will involve the isolation of immune cells from blood, then cultured and stimulated. The cells are contacted with an infection-sensitive cell line in contact. Maybe in the sample material available XMRV can multiply in the cultured cells and the number of detectable virus is thereby enlarged. If necessary, increased virus can then be detected using PCR.

To perform the test tube with a standard heparin-blood (10 ml) is required.

'Angebot: XMRV–Untersuchung bei CFS im deutschsprachigen Raum' English translation


VIRAL AND CELLULAR FACTORS GOVERNING EFFICIENT GENE DELIVERY

Looking for the source of XMRV in animals.

'VIRAL AND CELLULAR FACTORS GOVERNING EFFICIENT GENE DELIVERY'


Sandra Ruscetti/NCI - MOLECULAR BASIS FOR THE PATHOGENESIS OF MURINE RETROVIRUSES

  • Total Funding: $1,036,246

Friend spleen focus-forming virus (SFFV)causes a rapid, multi-stage leukemia in mice and provides a model for understanding how retroviral envelope proteins can have pathogenic consequences. Our previous studies have shown that the envelope protein of Friend SFFV interacts with and activates a unique receptor tyrosine kinase, sf-Stk, which is preferentially expressed in erythroid cells. This results in constitutive activation of signal transduction pathways for the growth, differentiation and survival of erythroid cells. Some of these erythroid cells become transformed due to SFFV integration, which results in activation of the myeloid transcription factor PU.1 and a block in erythroid cell differentiation. We showed that this block was due to activation of the hematopoietic phosphatase SHP-1, which in turn causes dephosphorylation of STAT1, blocking its ability to bind DNA and activate genes associated with differentiation. We further demonstrated that these SFFV-transformed erythroleukemia (SFFV-MEL) cells can metastasize to the bone marrow, where they are retained and subsequently cause meningeal leukemia, a common neurological complication of human leukemia.


Future work is aimed at better understanding the molecular basis for SFFV-induced meningeal leukemia and testing specific therapeutic strategies to block this retrovirus-induced multi-stage leukemia. In collaboration with Larry Keefer and members of his laboratory, we have shown that SFFV-MEL cells, but not normal hematopoietic cells, are killed by the nitric oxide prodrug JS-K. We demonstrated that JS-K induced necrosis and apoptosis in these cells by a caspase-dependent mechanism, causing DNA damage. We further demonstrated that JS-K treatment blocked the cell cycle by causing a block in the activation of the serine kinase Akt, leading to activation of the transcription factor FoxO3a, which in turn upregulated the cyclin-dependent kinase inhibitor p27. CDNB (1-chloro-2,4-dinitrobenzene), which contains an arylating ring analogous to that of JS-K without the diazeniumdiolate group necessary to release NO, was also able to kill SFFV-MEL cells by the same mechanisms, indicating that the arylating capability of JS-K, in addition to its ability to release NO, is a major contributor to the anticancer effects of this compound for SFFV-MEL cells. Studies are in progress to determine if JS-K can block the growth of SFFV-MEL cells in vivo. Several years ago we made the intriguing observation that SFFV can transform non-erythroid cells if it is co-expressed with sf-Stk. This results in transformation of fibroblasts in vitro and a variety of tumors in mice. We are currently utilizing the SFFV/sf-Stk-transformed fibroblasts to screen potential small molecule inhibitors for sf-Stk. We further showed that the human counterpart of sf-Stk, sf-RON, is expressed in certain types of human cancers, particularly ovarian and prostate cancers. Future work is aimed at determining whether sf-RON plays a causal role in these cancers and whether sf-RON, like sf-Stk, is activated by interacting with another protein, such as the envelope protein of a human retrovirus, to cause cancer.


As a second retroviral model system, we have been studying PVC-211 murine leukemia virus (MuLV), a variant of the erythroleukemia-inducing Friend MuLV that causes a rapid neurodegenerative disease in rodents. The diseased brains and spinal cords of PVC-211 MuLV-infected rats exhibit the spongiform pathology characteristic of some human neurodegenerative diseases such as HTLV-1-associated myelopathy/tropical spastic paraparesis and transmissible spongiform encephalopathy. We previously demonstrated that subtle changes in the envelope gene of the virus altered its host range from that of its leukemia-inducing parent Friend MuLV, enabling it to gain access to the central nervous system by infecting brain capillary endothelial cells (BCEC). By comparing BCEC from uninfected and PVC MuLV-infected rats, we found that virus-infected BCEC express high levels of inducible nitric oxide synthase and show evidence of NO production. We recently demonstrated that PVC-211 MuLV infection of BCEC in vivo results in the production of vascular endothelial cell growth factor and the chemokine MIP-1alpha, leading to vascular leakage and activation of microglia, the resident macrophages of the central nervous system. Further studies demonstrated that depletion of microglia from rat brains blocks neurodegeneration induced by PVC-211 MuLV and that treatment with antiserum to MIP-1alpha or splenectomy, both of which reduce the number of activated microglia in the brain, can delay disease, clearly demonstrating the importance of activated microglia in the development of PVC-211 MuLV-induced neurodegeneration. Current studies are focused on using pharmacological inerventions to block or mitigate PVC-211 MuLV-induced neurodegeneration.


We are currently using knowledge and reagents obtained from working with mouse retroviruses to study the xenotropic MuLV-related human retrovirus XMRV, which was recently discovered through an association with prostate cancer. In collaboration with the laboratories of Judy Mikovits and Frank Ruscetti, we were able to use antibodies developed against the envelope protein of SFFV to detect infectious XMRV in the blood cells and plasma of patients suffering from the neuroimmune disease chronic fatigue syndrome (CFS). We were further able to develop a seroconversion assay using cells expressing the SFFV envelope protein to detect antibodies against the virus in the plasma of CFS patients. We now plan to apply our knowledge of the pathogenesis of mouse retroviruses that cause cancer and neurological disease in rodents to study the molecular basis for similar diseases associated with XMRV. We are in the process of developing rodent models for determining the biological effects of XMRV in vivo, which if successful will provide a small animal model for preclinical testing of potential anti-XMRV drugs. In addition, we are testing both in vitro and in vivo the biological effects of the envelope protein of XMRV, which like its related SFFV counterpart may be responsible for the pathogenicity of XMRV.


'MOLECULAR BASIS FOR THE PATHOGENESIS OF MURINE RETROVIRUSES'


Further studies


SIMMONS, GRAHAM/NIH funded

DESCRIPTION (provided by applicant): Abstract Xenotropic murine leukemia virus-related virus (XMRV) is a newly identified gammaretrovirus linked to prostate cancer and chronic fatigue syndrome (CFS). Furthermore, other murine leukemia virus (MLV)-like sequences more closely related to polytropic MLV have also been reported to be associated with CFS. Since these viruses can be both detected in, and in the case of XMRV directly cultured from, leukocytes and plasma, they are highly likely to be transmissible by blood or blood products. Thus, sensitive, specific and reliable detection of the presence of XMRV, ideally by nucleic acid testing (NAT), is urgently required to further investigate this potentially serious threat to blood safety. Testing will be required initially to fully characterize the pathogenic potential of these viruses, together with the true prevalence in donor populations and the likelihood of transfusion-transmission (TT). These results will then determine whether large scale screening of the blood supply is warranted. Specific Aim 1: Detection of XMRV and other MLV-related viruses. Preliminary results, using validated clinical samples, have demonstrated that NAT detection of XMRV was, at best, variable. However, a 2 to 4 day delay, with samples held at 4oC prior to processing, allowed a more reliable detection of XMRV in the plasma fraction, by multiple laboratories. We hypothesize that regularly processed plasma is not the optimal medium for the detection of XMRV and other MLV-related viruses. Thus in aim 1 we will extend and further investigate these findings. We will perform detailed and extensive comparison of processing methods from multiple XMRV/MLV positive individuals. Plasma that becomes positive after a delay in processing will be treated with nucleases, proteases and detergents before and after ultracentrifugation, to determine whether viral nucleic acid is virion associated or present as free RNA/DNA. Specific Aim 2: Mechanisms to enhance nucleic acid testing. Although the optimization of the delayed processing of plasma will greatly enhance the potential of clinical and basic research studies, it is not likely to be practical for high-throughput screening of blood donors, where screening results are ideally required less than 24 hours after phlebotomy. Thus, we will investigate a multitude of techniques in order to achieve similar results to delayed processing, starting with those most likely to be easily and cheaply achieved in the blood collection setting.


PUBLIC HEALTH RELEVANCE: Xenotropic Murine Leukemia virus-Related Virus (XMRV) has been suggested to be involved in both prostate cancer and chronic fatigue syndrome, and furthermore has been demonstrated to be present in blood cells and plasma. Thus, there is real potential for XMRV transmission through blood or blood products and hence an urgent need for sensitive and specific detection assays in order to protect the blood supply. We plan to identify the most appropriate blood component for XMRV detection and will develop processing methods in order to maximize XMRV presence and hence detection within these components.

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