Method and compositions for use in diagnosing and...

Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving antigen-antibody binding – specific binding protein...

Reexamination Certificate

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C435S006120, C435S007100, C435S007400, C435S007920, C435S018000, C435S021000, C435S022000, C435S023000, C435S024000, C435S004000, C436S515000, C436S516000, C436S063000, C530S350000, C536S028100

Reexamination Certificate

active

06824988

ABSTRACT:

TECHNICAL FIELD
The field of this invention is chronic immune disease, particularly chronic fatigue syndrome and multiple sclerosis.
BACKGROUND OF THE INVENTION
Chronic immune diseases can be highly debilitating. Two such chronic immune diseases are multiple sclerosis and chronic fatigue syndrome.
Multiple sclerosis (MS) is a neurological illness of unknown etiology associated with attacks of focal or multifocal neurological dysfunction arising from lesions within the central nervous system (CNS). In America and Northern Europe, MS is the most common neurological disease, with prevalence rates estimated between 50-100 per 100,000 population. The onset of disease is most common in early adulthood. Recurrent attacks can occur over many years, with approximately 30 percent of the patients progressing to a severe form of the disease which can be fatal.
MS is pleomorphic in its presentation. The clinical manifestations are determined in part by the location of the foci of demyelination within the CNS. Classical features of the disease include impaired vision, nystagmus, dysarthria, ataxia and intention tremor, and weakness/paralysis of one or more limbs.
The most common form of the disease is episodic. Symptoms develop with subsequent recovery, then another attack occurs. In approximately 50 percent of all patients with MS, attacks become more frequent, usually with a worsening of symptomatology. In 30 percent of all patients, the disease develops into what is referred to as progressive/relapsing, the most severe form of the disease. In this state remissions are rare and patients frequently become wheelchair bound.
The characterization of MS disease activity (including diagnosis, determination of disease state, monitoring of disease progression, prediction of disease attacks, and the like), remains problematic. To aid the clinician, the only laboratory test available is testing the cerebrospinal fluid for oligoclonal bands, present in approximately 90 percent of all patients. Examination of the brain for demyelinating plaques, using magnetic resonance imaging (MRI) is useful but expensive, and is not warranted except in a small group of patients in which all other clinical and laboratory tests are negative. Furthermore, there is no diagnostic laboratory test to determine if a patient is having an “attack,” to monitor the progress of the “attack,” to determine if the patient is progressing to a more active form of the disease (i.e., progressive/relapsing), etc. Finally, there is no laboratory test available as a prognostic indicator and/or capable of monitoring the course of therapy. One commentator has summarized the situation as follows: “The need for reliable markers of disease activity in multiple sclerosis (MS) to better guide basic research, diagnosis, treatment, and monitoring therapy is well-recognized.” Laman et al., Mult. Scler. (Jun. 1998) 4:266-269.
Like MS, chronic fatigue syndrome (CFS) is an illness of unknown etiology. CFS is often associated with sudden onset, flu-like symptoms, debilitating fatigue, low-grade fever, myalgia and neurocognitive dysfunction. CFS patients typically display reduced Karnofsky performance scores (KPS). The Karnofsky performance test measures an individual's ability to function and carry on normal activities. Karnofsky scores range form zero for a nonfunctional or dead patient to 100 for a completely normal function.
Diagnosis of CFS remains one of exclusion. An accumulating body of evidence suggests that CFS is associated with disregulation of both humoral and cellular immunity, including mitogen response, reactivation of viruses, abnormal cytokine production, diminished natural killer cell function and changes in intermediary metabolites. It has been suggested that the clinical and immunological abnormalities observed in CFS might include defects in the double-stranded RNA (dsRNA)-dependent, interferon-inducible pathways, i.e., the 2′,5′-oligoadenylate (2-5A) synthetase/RNase L and p68 kinase (PKR) antiviral defense pathways (Suhadolnik et al., Clin. Infect. Dis. (1994) 18:S96-S104; Suhadolnik et al., In Vivo (1994 8:599-604. The 2-5A synthetase/RNase L pathway is part of the antiviral defense mechanism in mammalian cells; this pathway also has a role in the regulation of cell growth and differentiation (Lengyel, Ann. Review Biochem. (1982) 51:251-282; Sen et al., Adv. Virus Res. (1993) 42:57-102).
When activated by dsRNA, 2-5A synthetase converts ATP to 2′,5′-linked oligoadenylates with 5′-terminal phosphates. Biologically active 2-5A binds to and activates a latent endoribonuclease, RNase L, which hydrolyzes single-stranded viral and cellular RNA, primarily after UpNp sequences, thereby inhibiting protein synthesis.
Previous studies on the 2-5A synthetase/RNase L pathway in CFS revealed a statistically significant dysregulation in which the 2-5A synthetase is present predominantly in its activated form, bioactive 2-5A levels are elevated, and RNase L activity is upregulated (Suhadolnik et al., Clin. Infect. Dis., supra; Suhadolnik et al., In Vivo, supra). Expression of the serine-threonine kinase, PKR, is downregulated in CFS (Suhadolnik et al., In Vivo, supra). PKR controls initiation of protein translation through phosphorylation of eIF-2.
Despite these efforts, a clear cut molecular marker for CFS has not been identified. What is needed is a biochemical test, relying on an unambiguous molecular marker for CFS, which may form the basis of a definitive CFS diagnosis.
As the above discussion demonstrates, currently employed methods of diagnosing and/or characterizing MS or CFS disease activity in a subject are inadequate. As such, there is a continued need in the field to develop additional means for diagnosing and/or characterizing MS or CFS disease activity in a subject.
3. Relevant Literature
U.S. Pat. Nos. of interest include:5,766,859; 5,776,690; 5,830,668; 5,853,996; and 5,985,565. Other references of interest include: De Meirlier et al., Am. J. Med. (2000) 108:99-105; Komaroff, Am. J. Med. (2000) 108:69-171; Mashima et al., Oncogene (1999) 18:2423-2430; Mashima et al., Oncogene (1997) 14: 1007-1012; and Villa et al., J. Cell. Sci. (1998) 111: 713-722.
SUMMARY OF THE INVENTION
Methods are provided for diagnosing and/or characterizing chronic immune disease activity in a subject. In the subject methods, a sample is obtained from a subject suspected of having or known to have a chronic immune disease. The sample is then assayed for the presence of low molecular weight actin fragments. The assay results are used to diagnose the presence of chronic immune disease activity and/or characterize chronic immune disease activity in the subject, e.g. to confirm an initial chronic immune disease diagnosis, to determine the stage of the disease, to monitor disease progression, to predict disease attacks, and the like. Also provided by the subject invention are kits for practicing the methods.


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Komaroff (2000). “The biology of Chronic Fatigue Syndrome”Am. J. Med., 108: 169-171.
Mashima et al. (1997). “Actin cleavage by CPP-32/apopain during the development of apoptosis”Oncogene, vol. 14: 1007-1012.
Mashima et al. (1999). “Caspase-mediated cleavage of cytoskeletal actin plays a positive role in the process of morphological apoptosis”Oncogene, vol. 18: 2423-2430.
Meirleir et al. (2000). “A 37kDa 2-5A Binding Protein as a Potential Biochemical Marker for Chronic Fatigue Syndrome”Am. J. Med., vol. 108: 99-106.
Villa et al. (1998). “Calpain inhibitors, but not caspase inhibitors, prevent actin proceolysis and DNA fragmentation during apoptosis”J. Cell Sci., vol. 111(Pt

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