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1.
J Rheumatol ; 28(2): 257-65, 2001 Feb.
Article in English | MEDLINE | ID: mdl-11246659

ABSTRACT

OBJECTIVE: LJP 394 is a novel therapy under development for the treatment of systemic lupus erythematosus (SLE). We investigated the optimal LJP 394 dosing regimen required to maximally reduce serum dsDNA antibodies. We also evaluated the safety and tolerability of repeated doses of LJP 394 as well as the effects of therapy on SLE related disease activity and health related quality of life. METHODS: This was a multicenter, partially randomized, placebo controlled, double blind, dose-ranging trial. Study drug or placebo was administered at weekly, biweekly, or monthly intervals for a total of 17, 9, or 5 doses, respectively. Fifty-eight patients were randomly assigned to receive 1, 10, or 50 mg LJP 394 or placebo. After a 2 month pretreatment period, dosing visits continued for 16 weeks, after which there was a 2 month posttreatment period. RESULTS: The greatest reductions in mean dsDNA antibody titers were observed in the group of patients who received 50 mg LJP 394 weekly (38.1% and 37.1 % at Weeks 16 and 24, respectively). A reduction (29.3%) in dsDNA antibody titers was also observed at Week 24 in the group of patients who received 10 mg LJP 394 weekly. The frequencies of adverse events were comparable in the placebo and active treatment groups. CONCLUSION: This clinical trial, in which a large number of patients with SLE were treated with LJP 394, expanded the safety profile of LJP 394 and demonstrated its capacity to reduce dsDNA antibodies.


Subject(s)
Lupus Erythematosus, Systemic/drug therapy , Oligonucleotides/administration & dosage , Oligonucleotides/adverse effects , Adolescent , Adult , Aged , Antibodies/blood , DNA/immunology , Disability Evaluation , Female , Humans , Lupus Erythematosus, Systemic/immunology , Lupus Erythematosus, Systemic/physiopathology , Male , Middle Aged , Quality of Life , Surveys and Questionnaires
2.
Methods Mol Med ; 48: 39-54, 2001.
Article in English | MEDLINE | ID: mdl-21374404

ABSTRACT

Rapid drug susceptibility testing for Mycobacterium tuberculosis (Mtb) is imperative in an age when drug resistance is not rare and up to one-third of the world's population may be infected with this organism (1). A sensitive, PCRbased system to test mycobacterial antibiotic susceptibility is one approach to this problem. We reasoned that a quantitative PCR could detect the growth of bacilli by detecting an increase in the amount of mycobacterial DNA. Inclusion of an effective antibiotic in the culture media would prevent bacterial growth and concomitant increase in target DNA, thus distinguishing cultures which were susceptible to an antibiotic from those which were not.

4.
Rheum Dis Clin North Am ; 24(2): 227-36, 1998 May.
Article in English | MEDLINE | ID: mdl-9606756

ABSTRACT

Polymerase chain reaction and other DNA amplification techniques to identify elusive infections should prove to be an effective tool for the clinical and investigative rheumatologist. The capability to identify and characterize infectious organisms in fluids and tissue will enable early, specific, and potentially curative treatment. Similarly, the capability to exclude infection and differentiate postinfectious diseases will enable other therapies to control the inflammation. Understanding these molecular techniques will most certainly improve clinicians' effectiveness for diagnosis and care.


Subject(s)
Arthritis, Infectious/microbiology , DNA, Bacterial/isolation & purification , Polymerase Chain Reaction/methods , Whipple Disease/microbiology , Antigens, Bacterial/genetics , Arthritis, Infectious/diagnosis , DNA Primers , Humans , Whipple Disease/diagnosis
5.
J Clin Microbiol ; 35(5): 1161-5, 1997 May.
Article in English | MEDLINE | ID: mdl-9114400

ABSTRACT

Several problems remain before molecular biology-based techniques, such as PCR, are widely accepted for the detection of infectious agents. Among the most formidable of these problems are the inability of the tests to distinguish between viable and nonviable organisms. We approached this problem by using the fact that bacterial mRNA has an extremely short half-life, averaging only a few minutes. We reasoned that by targeting bacterial mRNA by a reverse transcriptase PCR (RT-PCR), a positive signal would indicate the presence of a recently viable organism. To test our hypothesis, we chose to target the mRNA coding for the ubiquitous 85B antigen of mycobacteria. After partially sequencing the gene coding for 85B, we developed primers that were specific for Mycobacterium tuberculosis. In a single-tube, nested, RT-PCR (STN RT-PCR), these primers detected fewer than 40 CFU in spiked sputum samples and as few as 12 CFU in clinical sputum specimens. The sensitivity of STN RT-PCR with smear-negative samples was as good as that of culture. The specificity was 100%. More importantly, when M. tuberculosis was cultured with and without 1 microgram of isoniazid per ml, this assay could distinguish between those cultures which contained the antibiotic and those which did not. Subcultures on Lowenstein-Jensen agar confirmed the viability assessments of the STN RT-PCR. Control experiments demonstrated that isoniazid did not inhibit the RT-PCR. In addition, when an IS6110-targeted, DNA PCR was used to examine the same samples, all samples though 13 days (the last sample) continued to be positive, irrespective of whether isoniazid was present, thereby demonstrating the superiority of an mRNA target in the detection of mycobacterial viability.


Subject(s)
Bacterial Typing Techniques , Mycobacterium tuberculosis/classification , Polymerase Chain Reaction/methods , Base Sequence , DNA, Bacterial/analysis , DNA, Bacterial/genetics , Molecular Sequence Data , Mycobacterium tuberculosis/isolation & purification
6.
Arthritis Rheum ; 37(5): 702-9, 1994 May.
Article in English | MEDLINE | ID: mdl-8185697

ABSTRACT

OBJECTIVE: To analyze synovial fluid (SF) for the presence of Neisseria gonorrhoeae DNA using the polymerase chain reaction (PCR). METHODS: We used a modified, nested PCR to detect the presence of N gonorrhoeae DNA in 41 samples of SF obtained from 10 patients with clinical gonococcal arthritis whose SF samples were sterile by culture and from 27 controls, including 11 patients with Reiter's syndrome. Results obtained using this method were compared with those obtained using the GEN-PROBE system, an RNA-DNA hybridization technique. RESULTS: With nested PCR, N gonorrhoeae DNA was detected in 11 of 14 SF samples obtained from patients with culture-negative clinical gonococcal arthritis but in none of the 11 SF samples from Reiter's syndrome patients. The specificity of this technique was 96.4%, with a sensitivity of 78.6%. The rate of false-positive results was 3.6%. The GEN-PROBE technique was unable to detect N gonorrhoeae ribosomal RNA in any of the samples. CONCLUSION: These findings demonstrate the potential utility of the PCR in confirming the clinical diagnosis of gonococcal arthritis as well as providing insight into the pathogenesis of this disorder in patients whose SF are sterile by standard culture techniques. PCR may also prove helpful in differentiating N gonorrhoeae arthritis from acute Reiter's syndrome.


Subject(s)
Arthritis, Infectious/diagnosis , Arthritis, Reactive/diagnosis , DNA, Bacterial/analysis , Gonorrhea/diagnosis , Neisseria gonorrhoeae/isolation & purification , Synovial Fluid/microbiology , Adolescent , Adult , Base Sequence , Diagnosis, Differential , Female , Humans , Male , Middle Aged , Molecular Sequence Data , Neisseria gonorrhoeae/genetics , Polymerase Chain Reaction , Predictive Value of Tests , Sensitivity and Specificity
7.
Arthritis Rheum ; 36(5): 665-75, 1993 May.
Article in English | MEDLINE | ID: mdl-8489545

ABSTRACT

OBJECTIVE: To analyze clinical fluids for the presence of Borrelia burgdorferi DNA using the polymerase chain reaction (PCR). METHODS: We utilized a modified, nested PCR to detect the presence of Borrelia DNA in 99 samples of serum, urine, cerebrospinal fluid (CSF), or synovial fluid obtained from 44 patients with various stages of Lyme disease and 47 control subjects. Primer specificity was corroborated by examining 2 DNA data banks, testing against DNA from other organisms, and confirming results with a second set of nested primers. RESULTS: Nested PCR was capable of detecting DNA from fewer than 10 organisms in 1 ml of fluid. The specificity of this technique was 96.4%, with a sensitivity of 76.7%. Although the specificity was uniformly high, the sensitivity was dependent upon the body fluid being tested: CSF 100%, urine 100%, synovial fluid 80%, and serum 59%. The rate of false-positive results was 3.6%. CONCLUSION: These data demonstrate the potential utility of PCR in confirming the clinical diagnosis of Lyme disease as well as providing insight into the pathogenesis of various stages of this disorder.


Subject(s)
Borrelia burgdorferi Group/isolation & purification , DNA, Bacterial/blood , Base Sequence , Borrelia burgdorferi Group/genetics , DNA, Bacterial/cerebrospinal fluid , DNA, Bacterial/genetics , DNA, Bacterial/urine , Gene Library , Humans , Molecular Sequence Data , Polymerase Chain Reaction , Sensitivity and Specificity
8.
Clin Immunol Immunopathol ; 55(3): 368-81, 1990 Jun.
Article in English | MEDLINE | ID: mdl-1692774

ABSTRACT

The DNA methylation inhibitor 5-azacytidine induces autoreactivity in cloned CD4+ T cells, but the functional consequences of this response are unknown. We now report that CD4+ T cells treated with 5-azacytidine respond to autologous antigen-presenting cells and induce autologous B cell differentiation without exogenous antigen or mitogen. This mechanism could play a role in some autoimmune diseases characterized by T cell DNA hypomethylation and polyclonal B cell activation.


Subject(s)
Azacitidine/pharmacology , B-Lymphocytes/immunology , CD4-Positive T-Lymphocytes/drug effects , Acecainide/pharmacology , Antibody Formation/physiology , Antigen-Presenting Cells/immunology , Antigens, CD/biosynthesis , Antigens, Differentiation/biosynthesis , CD4-Positive T-Lymphocytes/immunology , Cell Differentiation/immunology , DNA/drug effects , Histocompatibility Antigens/biosynthesis , Humans , Hydroxyurea/pharmacology , Integrin beta1 , Leukocyte Common Antigens , Methylation , Procainamide/pharmacology
9.
Mod Pathol ; 3(2): 186-91, 1990 Mar.
Article in English | MEDLINE | ID: mdl-2326250

ABSTRACT

Eighteen patients with lupus membranous nephritis were retrospective identified by reviewing the renal biopsy records of an active renal pathology service. Seven had minimally proliferative membranous nephropathy (World Health Organization (WHO) classes Va and b). Eleven had membranous nephropathy with superimposed changes of focal or diffuse proliferative glomerulonephritis (WHO Vc and d). After mean follow-up periods of 73 +/- 6 and 74 +/- 15 mo, respectively, one patient of seven from WHO Va and b and seven of 11 from WHO Vc and d reached end stage renal disease. The latter patients were distinguishable from the former only by the degree of superimposed proliferation on renal biopsy and not by blood pressure, antinuclear antibody, anti-double stranded DNA, or complement levels. These data stand in contrast to the widely held belief that lupus membranous nephropathy is relatively benign. The belatedness of this observation is partially due to imprecision in nosology for patients with lupus who have renal biopsies with "overlap" characteristics.


Subject(s)
Lupus Nephritis/pathology , Biopsy , Blood Pressure , Follow-Up Studies , Humans , Kidney/pathology , Kidney/physiopathology , Kidney Failure, Chronic/etiology , Lupus Nephritis/classification , Lupus Nephritis/physiopathology , Prognosis , Risk Factors , Serologic Tests
10.
J Clin Immunol ; 8(5): 362-71, 1988 Sep.
Article in English | MEDLINE | ID: mdl-2460488

ABSTRACT

To investigate the regulation of anti-DNA antibody production, we generated anti-DNA-specific suppressor cells by exposing normal human T cells and a small percentage of adherent cells to high concentrations of DNA. These cells suppressed the production of anti-DNA by both autologous peripheral blood mononuclear cells (PBMC) and allogeneic PBMC derived from systemic lupus erythematosus (SLE) patients. Anti-DNA production was suppressed significantly more than anti-RNA, antitetanus, or total immunoglobulin production. Specific suppression was enhanced by increasing the numbers of DNA-primed CD8+ cells and was obliterated by irradiation of the DNA-primed cells. In contrast to T cells from normal individuals, T cells obtained from two intensively studied SLE patients were unable to generate specific suppressor cells for anti-DNA production in both autologous and allogeneic test systems. Despite this defect, these patients were still capable of generating specific suppressor cells for antibody production directed against an exogenous antigen, tetanus toxoid.


Subject(s)
Antibodies, Antinuclear/biosynthesis , DNA/metabolism , T-Lymphocytes, Regulatory/metabolism , Cells, Cultured , DNA/radiation effects , Humans , In Vitro Techniques , Lupus Erythematosus, Systemic/metabolism , Macrophages/metabolism , Macrophages/radiation effects , RNA/metabolism , RNA/radiation effects , T-Lymphocytes, Regulatory/radiation effects
11.
Arthritis Rheum ; 28(4): 431-40, 1985 Apr.
Article in English | MEDLINE | ID: mdl-2580537

ABSTRACT

The spontaneous in vitro production of anti-DNA and anti-RNA by peripheral blood mononuclear cells (PBMC) from patients with systemic lupus erythematosus (SLE) and from normal subjects was evaluated employing sensitive solid-phase assays that are specific for these antibodies. PBMC from SLE patients produced more IgG anti-DNA and anti-RNA than did normal PBMC (P less than 0.01). In vitro production of IgG anti-DNA appeared to correlate with serum DNA bindings (r = 0.72, P less than 0.005). Similar amounts of IgM anti-DNA and anti-RNA were produced by both SLE and normal PBMC. However, IgM anti-DNA antibodies always appeared to be directed against determinants on denatured DNA. Only PBMC from SLE patients produced IgG antibodies to native DNA.


Subject(s)
DNA/immunology , Lupus Erythematosus, Systemic/immunology , Monocytes/immunology , RNA/immunology , Antibodies/analysis , Antibody Formation , Cells, Cultured , Humans , Immunoglobulin G/analysis , Immunoglobulin M/analysis , Radioimmunoassay
12.
Clin Immunol Immunopathol ; 27(3): 403-11, 1983 Jun.
Article in English | MEDLINE | ID: mdl-6603316

ABSTRACT

The titer of IgG antinucleoside antibodies in the sera of 162 individuals was determined by an enzyme-linked immunosorbent assay. The nucleosides used in the assay were adenosine, cytidine, guanosine, and thymine-riboside conjugated to human serum albumin. The specificity of IgG antinucleoside antibodies was indicated by appropriate reduction in antibody binding after solid-phase adsorptions of antibody with specific immobilized nucleoside conjugates. Disease-associated increases in serum IgG antibodies to cytidine and guanosine but not to adenosine or thymine-riboside occurred in patients with systemic lupus erythematosus (SLE). The epitope density of nucleosides in the conjugates and differences in the sensitivity of each nucleoside assay were not responsible for disease-associated IgG antinucleoside antibody responses. These findings support a possible pathogenic role for cytidine and guanosine as antigens or crossreactive antigenic determinants in some patients with SLE.


Subject(s)
Antibodies, Antinuclear/analysis , Antibody Specificity , Lupus Erythematosus, Systemic/immunology , Nucleosides/immunology , Adenosine/immunology , Binding Sites, Antibody , Cytidine/immunology , DNA, Single-Stranded/immunology , Guanosine/immunology , Humans , Immunoglobulin G/analysis
13.
J Rheumatol ; 9(4): 543-8, 1982.
Article in English | MEDLINE | ID: mdl-7131454

ABSTRACT

The results of a double-blind trial of pulse methylprednisolone in 9 patients with systemic lupus erythematosus and biopsy proven diffuse proliferative glomerulonephritis are reviewed. Patients were given 1 g of methylprednisolone or placebo intravenously, on 3 consecutive days, once a month, for 1 year. After 1 year, the methylprednisolone pulse group showed a significant improvement in serum creatinine whereas the placebo group showed none. More than 2 years after completing the study, renal function remained stable in the methylprednisolone group while the placebo group demonstrated significant deterioration in these values. Because of the limited number of patients involved, a concurrent control group was employed to confirm these results.


Subject(s)
Glomerulonephritis/drug therapy , Lupus Erythematosus, Systemic/drug therapy , Methylprednisolone/therapeutic use , Adult , Azathioprine/therapeutic use , Creatinine/blood , Double-Blind Method , Drug Therapy, Combination , Female , Humans , Infusions, Parenteral , Male , Methylprednisolone/adverse effects , Middle Aged
15.
Ann Intern Med ; 94(1): 21-6, 1981 Jan.
Article in English | MEDLINE | ID: mdl-7447218

ABSTRACT

Ten patients with rheumatoid arthritis unresponsive to conventional therapy participated in a double-blind cross-over trial in which they randomly received either a "pulse" or 1 g of methylprednisolone or placebo, intravenously, once a month for 6 months. Both the drug-first and placebo-first groups had the same mean American Rheumatism Association functional classification, 2.5. During the study patients on methylprednisolone "pulses," compared to placebo, showed significantly better mean tender-joint counts, walking times, and grip strength (p < 0.05). The drug-treated patients also had significantly lower levels of immune complexes (p < 0.01) and IgG (p < 0.01). Effects could still be measured an average of 2.9 +/- 0.4 months after the last dose of methylprednisolone. No significant side effects were noted during the therapy. Despite these findings, "pulse" methylprednisolone did not appear to significantly retard radiologic progression of the arthritis.


Subject(s)
Arthritis, Rheumatoid/drug therapy , Methylprednisolone/administration & dosage , Antigen-Antibody Complex/analysis , Arthritis, Rheumatoid/immunology , Arthritis, Rheumatoid/physiopathology , Double-Blind Method , Female , Humans , Immunoglobulin G/analysis , Infusions, Intravenous , Male , Middle Aged , Placebos
17.
J Rheumatol ; 5(3): 275-87, 1978.
Article in English | MEDLINE | ID: mdl-372523

ABSTRACT

Forty-seven SLE patients with severe renal disease characterized by renal biopsy documentation of diffuse proliferative or membranous glomerulonephritis or the nephrotic syndrome have been treated with azathioprine and prednisone in combination and followed for up to 12 years. Survivorship was 82% +/- 6% for five years and 74% +/- 8% for 10 years. There have been eight deaths and two patients have gone on hemodialysis. Five of the eight deaths are attributable to superinfection. Improvement in creatinine clearance was documented in 21 and decreased proteinuria in 35 of the patients. A therapeutic program, which included high dose corticosteroids initially, the combinations of azathioprine with corticosteroids chronically, and the rapid reduction in corticosteroid dosage to an alternate day schedule, appears to contribute to improved survivorship.


Subject(s)
Azathioprine/therapeutic use , Glomerulonephritis/drug therapy , Lupus Erythematosus, Systemic/drug therapy , Nephrotic Syndrome/drug therapy , Prednisone/therapeutic use , Adolescent , Adult , California , Clinical Trials as Topic , Drug Therapy, Combination , Female , Follow-Up Studies , Glomerulonephritis/complications , Humans , Lupus Erythematosus, Systemic/complications , Lupus Erythematosus, Systemic/mortality , Male , Middle Aged , Nephrotic Syndrome/complications
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