New Blood Markers Related to Chronic Fatigue Syndrome
May 1, 2000 6 minutes read
New Blood Markers Related to Chronic Fatigue Syndrome
abstract & commentary
Synopsis: De Meirleir and colleagues, through a series of intensely conducted studies, show that chronic fatigue syndrome patients often form an abnormal R-binding protein that likely is associated with dysregulation of 2-5A binding.
Source: De Meirleir K, et al. A 37 kDa 2-5A binding protein as a potential biochemical marker for chronic fatigue syndrome. Am J Med 2000;108:99-105.
Physicians are taught little about a cellular system designed to degrade residual single- stranded RNA. One such system uses the activation of an enzyme called RNase L. When this enzyme is activated through binding of a polynucleotide called 2-5-oligoadenylate (2-5A) it acts to degrade single-stranded RNA. The generation of 2-5A oligonucleotides begins with the priming of the cell by circulating interferons. Interferon signals the production of intracellular 2-5 synthetase. Next comes a crucial though poorly understood step. 2-5 synthetase requires the action of double-stranded RNA to become activated. Once activated and exposed to ATP, 2-5A is formed and then binds to RNase L, which then leads to the degradation of single-stranded RNA.
You might ask what is the true significance of this highly regulated system to degrade single-stranded RNA? The answer involves observations made primarily in in vitro analysis. Picornavirus, for example, clearly induces this system in vitro that regulates the replication of the virus. Blocks in the RNase L system can lead to increased viral replication. HIV, with all its intricacies, has a way of thwarting the RNase L system. In January 1999, Martinand and colleagues, including a coauthor in the current offering, showed that HIV induced an RNase L inhibitor that leads to upregulation of HIV replication.1 Other such inhibitors likely exist that allow unbridled replicaton of certain viruses.
Now enter chronic fatigue syndrome (CFS), for several years believed to be associated with a broad-based immune dysregulation. Even though CFS patients frequently have elevated antibody to various proteins of EB virus and human herpes virus 6 (HHV-6), these viruses play a causal role in CFS. In the current article, De Meirleir and colleagues, through a series of intensely conducted studies, show that CFS patients often form an abnormal R-binding protein that likely is associated with dysregulation of 2-5A binding.
De Meirleir has the largest CFS clinic in Europe, so he and his colleagues can study large numbers of patients. In the present study, they studied 57 patients who satisfied the CDC criteria for CFS. Controls were healthy subjects with fibromyalgia or depression. Another control group was 28 healthy "noncontact" controls recruited from the hospital and university staff. In this group were two persons who had "frequent contact with either CFS patients or their blood samples."
Identification of 2-5 binding activity used production of radiolabeled 2-5A that was incubated with protein extracts from peripheral blood mononuclear cells (PBMCs) and separated on an SDS gel using denaturing conditions. The proteins by size that were bound to 2-5A were visualized by autoradiography.
Both healthy subjects and CFS patients make normal 80 kDa and 40 kDa RNase L proteins. In 50 (88%) of CFS patients, however, there is a third protein band on SDS-PAGE seen around 37 kDa. It is present in only 28% of healthy controls and 38% of fibromyalgia patients, respectively. Two contact controls had elevated 37 kDa levels.
Each of the proteins suspected to be RNase Ls were shown to bind 2-5A, a necessary but not sufficient requirement of RNase Ls.
Analyzed another way, the ratio of the 37 kDa to the 80 kDa was equal to 0.5 in 72% of CFS patients, compared to only 1% of the healthy controls. These data emerge suggesting the potential for a sensitive, fairly specific, blood-based biochemical test.
Comment by Joseph F. John, MD
Those of us who care for CFS patients have known for several years of an abnormal R ding protein suggested from the work of Suhadolnik and colleagues.2 This new article from De Meirleir et al in Brussels (with basic science help from French workers) adds credence to the observation of defective RNase L proteins as applied to a larger group of CFS patients.
What does all this mean clinically? For starters, the credibility of CFS as a syndromic entity with cellular abnormalities certainly increases. Komaroff, a long-time CFS therapist and observer, in an accompanying editorial states clearly that there has been growing evidence for the immunological dysfunction in CFS.3 Now there is no doubt about some biochemical dysregulation.
So how should clinicians proceed with securing these data for patients suspected of having CFS? Few labs perform the assay for the low-molecular-weight RNase L. A lab known as RED provided the laboratory support for this study. The RNase L assay requires a pellet consisting of peripheral blood mononuclear cells that is well preserved in the cold. RED will process well-preserved PBMCs for RNase L proteins. Up to 88% of CFS patients will have PBMCs that produce the low-molecular-weight RNase L (DeMeirleir, personal communication) and the low-molecular-weight protein is expressed in few normal PBMCs.
Another lab here in the United States, known as ImmunoScience, also performs a set of tests for CFS using a slightly different method. Elevated levels of the 80 kDa RNase L are supposed to reflect the abnormality in CFS cells that not only produce a low-molecular-weight RNase L but also overproduce the 80 kDa protein.
There may be several reasons to monitor the 37 kDa protein in CFS patients. Therapies that decrease the level of the abnormal protein may be associated with clinical improvement. Insurance companies, which for years have balked at providing disability for CFS patients, may welcome a biochemical marker for the disease. Finally, patients themselves will gain peace of mind knowing that their multisystem symptomatology is associated with some cellular abnormality.
The biology of CFS now suggests that a cellular abnormality in RNA metabolism may be associated with the disease.3 Work in vitro with picornavirus and reovirus has found that an intact R system as stimulated by interferon results in effective degradation of single-stranded viral RNA. Interestingly, HIV RNA degradation is defective in HIV-infected patients due to an inhibitor of R that serves to allow upregulation of HIV and increased viral replication. Other viral systems have not been well studied.
What is far from clear is whether the R system functions to degrade host cell RNA. If the differential effect of R on host vs. viral RNA can be demonstrated, we will have a better appreciation for the R defect in CFS patients. If the R system serves primarily to degrade viral RNA, we must continue to search for a viral cause of CFS.
In process is a clinical trial of an interferon inducer known as polyI:polyC (Ampligen) that has enrolled more than 125 patients. The Ampligen 516 trial is a double-blinded, placebo-controlled format that includes analysis of the RNase L system in CFS patients treated with Ampligen or placebo. We all eagerly await the results of that trial.
References
1. Martinand C, et al. RNase L inhibitor is induced during HIV type 1 infection and down regulates the 2-5A/RNase L pathway in human T cells. J Virol 1999; 73:290-296.
2. Suhadolnik RJ, et al. Biochemical evidence for a novel low molecular weight 2-5A-dependent RNase L in chronic fatigue syndrome. J Interferon Cytokine Res 1997;17:377-385.
3. Komaroff AL. The biology of chronic fatigue syndrome. Am J Med 2000;108:169-171.
The R system provides for metabolism of what product?
a. DNA
b. Single-stranded RNA
c. Double-stranded RNA
d. 2’5’ oligoadenylate (2-5A)
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