Lynell W. Klassen

Treatment of Rheumatoid Arthritis with Methotrexate Alone, Sulfasalazine and Hydroxychloroquine, or a Combination of All Three Medications

BACKGROUND Rheumatoid arthritis is a common disease that causes substantial morbidity and mortality. The responses of patients with rheumatoid arthritis to treatment with a single so-called disease-modifying drug, such as methotrexate, are often suboptimal. Despite limited data, many patients are treated with combinations of these drugs. METHODS We enrolled 102 patients with rheumatoid arthritis and poor responses to at least one disease-modifying drug in a two-year, double-blind, randomized study of treatment with methotrexate alone (7.5 to 17.5 mg per week), the combination of sulfasalazine (500 mg twice daily) and hydroxychloroquine (200 mg twice daily), or all three drugs. The dose of methotrexate was adjusted in an attempt to achieve remission in all patients. The primary and point of the study was the successful completion of two years of treatment with 50 percent improvement in composite symptoms of arthritis and no evidence of drug toxicity. RESULTS Fifty of the 102 patients had 50 percent improvement at nine months and maintained at least that degree of improvement for two years without evidence of major drug toxicity. Among them were 24 of 31 patients treated with all three drugs (77 percent), 12 of 36 patients treated with methotrexate alone (33 percent, P < 0.001 for the comparison with the three-drug group), and 14 of 35 patients treated with sulfasalazine and hydroxychloroquine (40 percent), P = 0.003 for the comparison with the three-drug group). Seven patients in the methotrexate group and three patients in each of the other two groups discontinued treatment because of drug toxicity. CONCLUSIONS In patients with rheumatoid arthritis, combination therapy with methotrexate, sulfasalazine, and hydroxychloroquine is more effective than either methotrexate alone or a combination of sulfasalazine, and hydroxychloroquine.

Treatment of early seropositive rheumatoid arthritis: A two-year, double-blind comparison of minocycline and hydroxychloroquine

OBJECTIVE To compare the efficacy of minocycline with that of a conventional disease-modifying antirheumatic drug (DMARD), hydroxychloroquine, in patients with early seropositive rheumatoid arthritis (RA). METHODS Sixty patients with seropositive RA of <1 years duration who had not been previously treated with DMARDs were randomized to receive minocycline, 100 mg twice per day, or hydroxychloroquine, 200 mg twice per day, in a 2-year, double-blind protocol. All patients also received low-dose prednisone. The primary end points of the study were 1) the percentage of patients with an American College of Rheumatology (ACR) 50% improvement (ACR50) response at 2 years, and 2) the dosage of prednisone at 2 years. RESULTS Minocycline-treated patients were more likely to achieve an ACR50 response at 2 years compared with hydroxychloroquine-treated patients (60% compared with 33%, respectively; P = 0.04). Minocycline-treated patients were also receiving less prednisone at 2 years compared with the hydroxychloroquine group (mean 0.81 mg/day compared with 3.21 mg/day, respectively; P < 0.01). In addition, patients treated with minocycline were more likely to have been completely tapered off prednisone (P = 0.03). Trends favoring the minocycline treatment group were seen when outcomes were assessed according to components of the ACR core criteria set, with the differences reaching statistical significance for patients global assessment of disease activity (P = 0.004). CONCLUSION Minocycline is an effective DMARD in patients with early seropositive RA. Patients treated with minocycline were more likely to achieve an ACR50 response and did so while receiving less prednisone. In addition, minocycline-treated patients were more likely to have discontinued treatment with prednisone at 2 years.

Treatment of early seropositive rheumatoid arthritis with minocycline: Four‐year followup of a double‐blind, placebo‐controlled trial

OBJECTIVE Rheumatoid arthritis (RA) causes substantial morbidity and mortality, and current treatments are suboptimal. Recent studies have demonstrated the short-term efficacy of minocycline in the treatment of patients with early RA. This study was undertaken to compare patients treated with conventional therapy in the early phase of their RA and those treated with minocycline, after 4 years of followup. METHODS Forty-six patients with seropositive RA of <1 years duration had been enrolled in a double-blind study of minocycline (100 mg twice daily) versus placebo. After the blinded portion of the study (3-6 months, depending upon response), all patients were treated with conventional therapy. This report compares those patients randomized to receive placebo for 3 months and then conventional therapy for the duration of 4 years versus those originally randomized to receive minocycline. RESULTS Twenty of the 23 original minocycline-treated patients and 18 of the 23 original placebo-treated patients were available for followup (mean 4 years). At followup, RA was in remission (American College of Rheumatology criteria) without disease-modifying antirheumatic drug (DMARD) or steroid therapy in 8 of the patients originally treated with minocycline compared with 1 patient in the placebo group (P = 0.02). Ten patients in the minocycline group versus 16 in the original placebo group currently require DMARD therapy (P = 0.02). CONCLUSION Among patients with seropositive RA, remissions are more frequent and the need for DMARD therapy is less in those treated early in the disease course with minocycline compared with those treated with conventional therapy delayed by an average of only 3 months. Minocycline appears to be an effective therapy for early RA; further investigation into its mechanism of action is needed.

HLA-DRB1 typing in rheumatoid arthritis: predicting response to specific treatments

OBJECTIVE To determine the predictive value of shared epitope alleles for response to treatment in patients with rheumatoid arthritis. METHODS Patients from our previously published triple DMARD study were tested for the presence of shared epitope alleles (DRB1 *0401,0404/0408, 0405,0101, 1001,and 1402). Patients who were shared epitope positive were then compared with those who were negative to see if there was a differential effect on therapeutic response. RESULTS Shared epitope positive patients were much more likely to achieve a 50% response if treated with methotrexate-sulphasalazine-hydroxychloroquine compared with methotrexate alone (94% responders versus 32%, p<0.0001). In contrast shared epitope negative patients did equally well regardless of treatment (88% responders for methotrexate-sulphasalazine-hydroxychloroquine versus 83% for methotrexate). Additionally, a trend toward an inverse relation of the gene dose was seen for response to methotrexate treatment (p=0.05). CONCLUSIONS These data suggest that determining shared epitope status may provide clinical information useful in selecting among treatment options.

Detection of circulating antibodies to malondialdehyde-acetaldehyde adducts in ethanol-fed rats

BACKGROUND & AIMS Malondialdehyde and acetaldehyde react together with proteins and form hybrid protein conjugates designated as MAA adducts, which have been detected in livers of ethanol-fed rats. The aim of this study was to examine the immune response to MAA adducts and other aldehyde adducts during long-term ethanol exposure. METHODS Rats were pair-fed for 7 months with a liquid diet containing either ethanol or isocaloric carbohydrate. Circulating antibody titers against MAA adducts and acetaldehyde adducts were measured and characterized in these animals. RESULTS A significant increase in antibody titers against MAA-adducted proteins was observed in the ethanol-fed animals. Competitive inhibitions of antibody binding indicated that the circulating antibodies against MAA-modified proteins in the ethanol-fed rats recognized mainly a specific, chemically defined MAA epitope. Antibody titers to reduced and nonreduced acetaldehyde adducts were very low, and no significant differences were observed between ethanol-fed and control animals. Significant plasma immunoreactivity to not only MAA-adducted but also unmodified rat liver proteins (cytosol, microsomes, and especially plasma membrane) were also observed in the ethanol-fed rats. CONCLUSIONS Long-term ethanol feeding generates circulating antibodies not only against MAA epitopes but possibly also against unmodified, native (self) protein epitopes, suggesting that MAA adducts could trigger harmful autoimmune responses.

Association of malondialdehyde-acetaldehyde (MAA) adducted proteins with atherosclerotic-induced vascular inflammatory injury

Atherosclerosis is a vascular injury characterized by elevated tissue levels of tumor necrosis factor-alpha (TNF-alpha), increased expression of endothelial cell adhesion molecules, and vascular wall inflammatory cell infiltration. Foam cells are associated with atherosclerotic plaque material, and low density lipoprotein (LDL) is a lipid component of foam cells. Malondialdehyde (MDA) is an oxidative product of unsaturated fatty acids and is also present in atherosclerotic lesions. MDA-modified (adducted) proteins, including MDA-modified LDL, are present in atherosclerotic human vascular tissue. Acetaldehyde (AA) is the major metabolic product of ethanol oxidation. Both MDA and AA are highly reactive aldehydes and will combine with proteins to produce an antigenically distinct protein adduct, termed the MAA adduct. This study demonstrates that proteins modified in the presence of high concentrations of MDA can produce MAA-modified proteins in vitro. In addition, MAA adducted proteins are capable of inducing rat heart endothelial cell cultures (rHEC) to produce and release TNF-alpha, and cause rHEC upregulation of endothelial adhesion molecule expression, including ICAM-1. These adhesion molecules are required for circulating inflammatory cells to adhere to endothelium which allows inflammatory cell tissue infiltration. Additionally, MAA modified proteins were defected in human atherosclerotic aortic vascular tissue but not in normal aortic tissue. Since atherosclerosis is associated with an inflammatory vascular injury characterized by elevated tissue TNF-alpha concentrations and inflammatory cell infiltration, these data suggest that MAA-adducted proteins may be formed in atherosclerotic plaque material and may be involved in the inflammatory reaction that occurs in atherosclerosis. These data further suggest that previous studies demonstrating MDA modified protein in atherosclerotic plaque may in fact have MAA modified proteins associated with them.

Lipopolysaccharide is a cofactor for malondialdehyde-acetaldehyde adduct-mediated cytokine/chemokine release by rat sinusoidal liver endothelial and kupffer cells

BACKGROUND The nonparenchymal cells of the liver have been suggested to play a significant role in the inflammatory processes observed in the development and/or progression of alcoholic liver disease. Our laboratories have shown that malondialdehyde-acetaldehyde (MAA)-modified proteins can induce immune responses, cytokine/chemokine secretion, and antigen processing and presentation by liver sinusoidal endothelial cells (SECs). Another molecule that has been shown to induce similar types of responses in Kupffer cells (KCs) is lipopolysaccharide (LPS). Because these materials induce similar responses, it was the purpose of this study to investigate the relationship between LPS and MAA-modified proteins in the development of proinflammatory responses by SECs and KCs. METHODS For these studies, SECs and KCs were isolated from chow-fed, pair-fed, and ethanol-fed rats. Cells were stimulated with media alone, bovine serum albumin (Alb), or MAA-modified Alb (MAA-Alb) in the presence or absence of LPS 1 ng/ml, and the supernatants were assayed by enzyme-linked immunosorbent assay for tumor necrosis factor alpha, macrophage chemotactic protein 1, and macrophage inhibitory protein. RESULTS All three cytokines/chemokines were 3 to 5 times higher when SECs or KCs were stimulated by MAA-Alb in the presence of LPS, in contrast to cells stimulated with Alb or media in the presence of LPS. Chronic ethanol consumption (6 weeks) had variable effects on the secretion of these cytokines/chemokines but in general did not alter the increased secretion in response to MAA-Alb in the presence of LPS. CONCLUSIONS These studies strongly suggest that the sensitization of SECs and KCs by LPS plays a significant role in the development and/or progression of alcoholic liver disease, and the subsequent activation by MAA-modified proteins may be a mechanism by which proinflammatory processes are initiated.

Malondialdehyde–acetaldehyde adduct is the dominant epitope after MDA modification of proteins in atherosclerosis

Antibodies to malondialdehyde (MDA)-modified macromolecules (adducts) have been detected in the serum of patients with atherosclerosis and correlate with the progression of this disease. However, the epitope and its formation have not been characterized. Studies have shown that excess MDA can be degraded to acetaldehyde, which combines with proteins to from a stable dihydropyridine adduct. To investigate, mice were immunized with MDA adducts in the absence of adjuvant and showed an increase in antibodies to MDA adducts and the carrier protein as the concentration of MDA was increased. In fact, a number of the commercially available antibodies to MDA-modified proteins were able to be inhibited by a chemical analogue, hexyl-MAA. Also, MDA-MAA adducts were detected in the serum and aortic tissue of JCR diabetic/atherosclerotic rats. These studies determined that commercially available antibodies to MDA predominantly react with the MAA adduct and are present in the JCR model of atherosclerosis in both the serum and the aortic tissue. Therefore, the immune response to MDA-modified proteins is most probably to the dihydropyridine structure (predominant epitope in MAA), which suggests that MAA adducts may play a role in the development and/or progression of atherosclerosis.

Malondialdehyde-Acetaldehyde Adducts and Anti–Malondialdehyde-Acetaldehyde Antibodies in Rheumatoid Arthritis

Malondialdehyde‐acetaldehyde (MAA) adducts are a product of oxidative stress associated with tolerance loss in several disease states. This study was undertaken to investigate the presence of MAA adducts and circulating anti‐MAA antibodies in patients with rheumatoid arthritis (RA).

IL-6 and its receptors in coronary artery disease and acute myocardial infarction.

Biomarkers such as interleukin-6 (IL-6), soluble interleukin-6 receptor (sIL-6R), and high sensitive C-reactive protein (hsCRP) have been reported to be elevated in acute myocardial infarction (AMI). The aim of this study is to determine the relationship between these markers during AMI, as well as their relationship to clinical parameters in an effort to discern their predictive potential in cardiac events. Serum was collected from 73 patients with; AMI, stable coronary artery disease (CAD), and controls during cardiac catheterization. Biomarker levels were determined and correlated with clinical data. IL-6 (11.75pg/ml, P<0.05) and sIL-6R (41,340pg/ml, P=0.05) were elevated in AMI compared with CAD and controls. At presentation, hsCRP was elevated in AMI patients (4.69mg/L) compared to controls (2.69mg/L, P<0.05); however, there was a significant decrease in hsCRP between AMI (4.69mg/L) and CAD patients (7.4mg/L, P<0.05). After 24h post-AMI hsCRP levels were increased compared to stable CAD (60.46mg/L, P<0.05) and were preceded by increased IL-6 at presentation. Soluble Gp130 (sGp130) showed no significant change between AMI, CAD, and control patients. However, sGp130 positively correlated with peak troponin in AMI (R=0.587, P<0.01), and negatively correlated with previous AMI (R=-0.382, P<0.05). Circulating monocyte mRNA expression isolated from selected AMI patients showed an increase in IL-6 mRNA (5.28-fold, P<0.01) and a decrease in both IL-6R (0.374-fold, P<0.01) and sGp130 mRNA (0.38-fold, P<0.01) as compared to CAD and controls. Results demonstrate that IL-6 and sIL-6R are associated with AMI and cardiac injury. These data support the hypothesis that trans-IL-6 receptor binding may alter intracellular signaling, and blocking of IL-6 receptor binding may be pathogenic in AMI. These data may be predictive of mechanism(s) by which plaques become unstable and rupture.