Experiences with dose finding in patients in early drug development: the use of biomarkers in early decision making.

With the increasing cost and complexity of drug development, biomarkers will play an increasing role in the early phases. Biomarkers can be classified into target, mechanistic, or outcome with varying degrees of linkage to disease or treatment effect. They can be used to determine proof of concept by characterising the efficacy or safety profiles, or determining differentiation from any competitor drugs. PK/PD modelling of biomarker data for novel and marketed compounds can be used to predict outpatient dose response. Subsequent simulations may replace or reduce the size and cost of larger phase 2b outpatient studies. Two examples of biomarkers and PK/PD modelling used to characterise dose response are presented. Penile plethysmography (RigiScan Plus) in male erectile dysfunction and phenylephrine challenge urethral pressure in benign prostatic hyperplasia are used to reduce time and cost to reach major exploratory development decision points in these indications.

Analysis of changes in acute-phase plasma proteins in an acute inflammatory response and in rheumatoid arthritis using two-dimensional gel electrophoresis.

Two-dimensional (2-D) gel analysis was used to examine differences in the levels of 19 plasma proteins: before and after an acute inflammatory reaction (parenteral typhoid vaccination) in normal subjects, between rheumatoid arthritis (RA) patients and normals and in RA patients treated with tenidap (120 mg) and piroxicam (20 mg). Typhoid vaccination increased levels of SAA, haptoglobin alpha1, haptoglobin alpha2, haptoglobin beta and alpha1-anti-chymotrypsin but decreased transthyretin and apolipoprotein E. In RA patients, serum amyloid A (SAA), haptoglobin alpha2, haptoglobin beta, alpha1-antichymotrypsin and C3 proactivator levels were elevated while apolipoprotein A-I, apolipoprotein A-IV, transthyretin, Gc-globulin, alpha2-HS glycoprotein, alpha2-macroglobulin and alpha1-B glycoprotein levels were decreased, compared to normals. Compared to piroxicam, tenidap lowered levels of alpha1-antiprotease and SAA but raised the levels of transthyretin, Gc-globulin, alpha2-HS-glycoprotein and alpha2-macroglobulin in RA patients. C-reactive protein (CRP) could not be quantified on 2-D gels but, when measured by rate nephelometry, levels were reduced after treatment with tenidap compared to piroxicam. The general pattern of the acute phase protein response to an acute inflammatory response to typhoid vaccination is similar to that in the chronic inflammatory condition, RA. The impact of tenidap on both positive and negative acute-phase proteins in RA patients could clearly be distinguished from that of piroxicam.

Effect of tenidap on metalloproteinase gene expression in rheumatoid arthritis: a synovial biopsy study.

This study evaluated measures of metalloproteinase gene expression in synovial biopsies as a means of differentiating the activities of two antirheumatic therapies: piroxicam and tenidap. Synovial biopsies and quantitative in situ hybridization for stromelysin (STR), collagenase (COL), tissue inhibitor of metalloproteinase-1 (TIMP), and actin mRNA were performed in a subset of patients with rheumatoid arthritis in a larger doubleblind randomized crossover trial comparing 120 mg/day of tenidap for 6 weeks with 20 mg/day of piroxicam for 6 weeks. There were no consistent differences between tenidap and piroxicam on COL or TIMP mRNA expression, but STR mRNA was significantly lower after tenidap compared with piroxicam (p = 0.037). There were no significant associations between measures of local or systemic clinical activity and STR, COL, or TIMP gene expression. However, changes in STR gene expression were significantly correlated with changes in serum C-reactive protein (p = 0.016). Because tenidap and piroxicam are both potent inhibitors of prostaglandin production, the effect of tenidap on STR gene expression may be due to its additional cytokine modulating activity. Clinical trials with data from synovial biopsies may be useful in evaluating the potential for disease-modifying effects of antirheumatic drugs.

Steroid sparing activity of tenidap in patients with polymyalgia rheumatica: a multicenter double blind randomized placebo controlled study.

OBJECTIVE: To determine whether tenidap treatment would allow reduction or replacement of systemic corticosteroid treatment in patients with polymyalgia rheumatica (PMR). METHODS: A 15-week double blind, randomized, multicenter, placebo-controlled study of tenidap sodium (120 mg/day) in patients with symptomatically controlled PMR receiving 10 mg/day prednisone was conducted. After receiving study drug for 3 weeks, prednisone dose was reduced by 2.5 mg/day every 3 weeks. The lowest clinically effective dose of prednisone was recorded as 10, 7.5, 5, 2.5 or 0 mg/day. RESULTS: Thirty-two patients were randomized to tenidap or placebo. As prednisone was reduced more placebo patients experienced an exacerbation of PMR symptoms, elevation of erythrocyte sedimentation rate and increased serum C-reactive protein. Twice as many placebo patients (10 of 16) as tenidap patients (5 of 16) discontinued due to lack of efficacy. The lowest effective dose of prednisone could be determined in 27 of the 32 patients, 11 receiving tenidap and 16 placebo. A significantly (p = 0.027) greater proportion of patients receiving tenidap (5 of 11) than placebo (1 of 16) were able to discontinue prednisone without experiencing a symptomatic flare. CONCLUSION: As prednisone was reduced, symptoms of PMR were controlled better by tenidap than by placebo. Forty-five percent of evaluable patients receiving tenidap were able to discontinue prednisone without a disease flare compared to 6% for placebo.

Products of arachidonic acid metabolism and the effects of cyclooxygenase inhibition on ongoing cutaneous allergic reactions in human beings.

BACKGROUND: There have been conflicting reports about the effects of inhibition of arachidonic acid metabolism on early- and late-phase cutaneous reactions. We re-examined this question with a unique nonsteroidal antiinflammatory drug, tenidap sodium. Tenidap sodium has been demonstrated in in vitro studies to inhibit cyclooxygenase, lipoxygenase, and cytokine production (interleukin-1, interleukin-6, tumor necrosis factor-alpha). METHODS: In a double-blind, randomized, crossover study, seven pollen-sensitive subjects ingested tenidap (120 mg, by mouth, daily) and placebo for 9 days with a 3-week washout period between treatments. On the eighth day they underwent allergen skin testing, measurable for up to 12 hours, and on the ninth day they underwent 5-hour skin chamber exposures to allergen and buffer. Chamber fluids were analyzed for cellular content, neutrophil granule protein release, cyclooxygenase and lipoxygenase arachidonic acid metabolites, histamine, and tryptase. RESULTS: Tenidap did significantly inhibit cyclooxygenase metabolites at both antigen and buffer sites but had no effect on histamine, tryptase, lipoxygenase metabolites, or granulocyte infiltration. Neutrophil granule release of lactoferrin was lower at the antigen site during tenidap administration, but there was no reduction of elastase release. Prostaglandin E2 and leukotriene E4 increased significantly at antigen sites compared with buffer sites during placebo administration and were the most prominent arachidonic acid metabolites detected. CONCLUSION: Tenidap, despite inhibiting cyclooxygenase release at antigen sites, had no effect on skin test responses to antigen or on antigen-induced mediator release or granulocyte infiltration. We conclude that cyclooxygenase metabolites are not important in the development of an allergic cutaneous inflammatory response.

Rheumatoid arthritis treated with tenidap and piroxicam. Clinical associations with cytokine modulation by tenidap.

OBJECTIVE: To compare the effects of tenidap and piroxicam on acute-phase protein and cytokine levels in the blood of rheumatoid arthritis (RA) patients and to explore their associations with clinical disease activity. METHODS: A double-blind, randomized, crossover trial in 49 patients with active RA compared 6 weeks of treatment with tenidap (120 mg/day) versus 6 weeks of treatment with piroxicam (20 mg/day). RESULTS: Median values for C-reactive protein (CRP), Westergren erythrocyte sedimentation rate (ESR), serum amyloid A (SAA) protein, and interleukin-6 (IL-6) were significantly lower after tenidap treatment compared with piroxicam treatment, even in the presence of stable background treatment with prednisone, methotrexate, or prednisone plus methotrexate. The median within-patient treatment differences (after tenidap minus after piroxicam) in the CRP, ESR, SAA, and IL-6 values were -1.7 mg/dl, -10.0 mm/hour, -22.0 micrograms/ml, and -3.7 pg/ml, respectively, and represent -60.4%, -17.7%, -35.5%, and -26.1% of the respective baseline levels. IL-6 levels were positively correlated with CRP and SAA. Plasma IL-1 beta was generally below the level of detection. Tumor necrosis factor alpha levels were similar after tenidap and after piroxicam. Treatment differences for 4 of 7 clinical parameters favored tenidap, but did not reach statistical significance. IL-6, CRP, and ESR were significantly correlated with clinical treatment differences. Tenidap and piroxicam toleration were similar, although tenidap-treated patients exhibited a reversible increase in urinary protein excretion. CONCLUSION: Tenidap was differentiated from piroxicam by lower levels of acute-phase proteins, ESR, and IL-6 after tenidap treatment. These treatment differences were significantly correlated with clinical parameters.

Elevated cerebrospinal fluid levels of substance P in patients with the fibromyalgia syndrome.

OBJECTIVE: To measure, and seek clinical correlates with, levels of substance P (SP) in the cerebrospinal fluid (CSF) of fibromyalgia syndrome (FMS) patients. METHODS: CSF from 32 FMS patients and 30 normal control subjects was tested for SP by radioimmunoassay. Clinical measures included tender point examination and standardized questionnaires. RESULTS: CSF SP levels were 3-fold higher in FMS patients than in normal controls (P < 0.001), but they correlated only weakly with tenderness found on examination. CONCLUSION: SP is significantly elevated in FMS CSF, but other abnormalities must exist in FMS to more fully explain the symptoms.

Gene expression (collagenase, tissue inhibitor of metalloproteinases, complement, and HLA-DR) in rheumatoid arthritis and osteoarthritis synovium. Quantitative analysis and effect of intraarticular corticosteroids.

In situ hybridization was used to localize and quantify gene expression in rheumatoid arthritis (RA) and osteoarthritis (OA) synovial tissue. Collagenase, tissue inhibitor of metalloproteinases (TIMP), HLA-DR, and complement (C2 and C3) gene expression was studied in synovial tissue from 23 patients with RA, OA, or other inflammatory arthropathies. Gene expression was highly compartmentalized: Collagenase, TIMP, and C2 messenger RNA (mRNA) were localized primarily to the synovial lining layer; HLA-DR mRNA was prominent in the lining and in some sublining lymphoid aggregates; the C3 probe hybridized only to sublining lymphoid aggregates. Relative mRNA levels were quantified using computer-assisted image analysis. There was significantly more collagenase, C2, C3, and HLA-DR mRNA in RA compared with OA patients. However, TIMP mRNA levels were similar in RA and OA. Expression of collagenase, TIMP, C2, C3, and HLA-DR genes correlated with the degree of synovial inflammation. The effect of intraarticular corticosteroid injection on synovial tissue gene expression was studied using serial percutaneous synovial biopsy samples from the knees of 3 RA patients. Joints were biopsied, injected with triamcinolone, and rebiopsied 1-2 weeks later. Histologic inflammation scores were lower in posttreatment synovia. Collagenase and TIMP mRNA, although abundant in presteroid samples, were nearly undetectable in post-steroid tissues. HLA-DR mRNA levels also were significantly decreased. C2 and C3 hybridization significantly decreased in 2 of 3 patients and 1 of 3 patients, respectively. Hence, clinical response to intraarticular steroid therapy was accompanied by histologic improvement and decreased expression of genes that play a role in articular destruction.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of gold sodium thiomalate and tenidap sodium (CP-66,248-2) on a model of macrophage differentiation using HL-60 cells.

We studied the effects of gold sodium thiomalate (GST) and a new antirheumatic drug, tenidap sodium ([Z]-5-chloro-2,3-dihydro-3-[hydroxy-2-thienylmethylene]-2-oxo-1H- indole-1-carboxamide, sodium salt), previously known as CP-66,248-2, in a model system of macrophage differentiation using a myelomonocytic cell line. HL-60 cells can be stimulated by vitamin D3 to differentiate along a monocytic pathway. Monocytic HL-60 cells express CD14 (Leu-M3), a macrophage surface marker, and develop the capacity to produce the second complement component (C2) in response to stimulation with cytokines such as gamma-interferon. The effects of GST and tenidap sodium were compared with the effects of dexamethasone and a variety of nonsteroidal antiinflammatory drugs in this model system. We found that GST inhibited the capacity of HL-60 cells to produce C2 but did not inhibit the expression of CD14. Tenidap sodium inhibited C2 production as well as CD14 expression, and it partially reversed the decrease in 3H-thymidine incorporation by HL-60 cells, which accompanies monocytic differentiation. At concentrations that inhibited C2 production by HL-60 cells, tenidap sodium did not inhibit C2 production by monocytes. Neither dexamethasone nor the other nonsteroidal antiinflammatory drugs tested possessed these activities. Thus, both GST and tenidap inhibit markers of monocytic differentiation in HL-60 cells, and this activity may relate to their antirheumatic activities.

Linear scleroderma: a response to neurologic injury? Report and literature review.

A 28-year-old white man presented with neurologic symptoms and skin changes. Subsequent evaluation led to the diagnosis of transverse myelitis of the cervical spine (C8) and linear scleroderma. The progression of neurologic abnormalities prompted treatment with corticosteroids. Neurologic symptoms diminished and the progression of linear skin lesions halted. A review of the literature uncovered considerable evidence for underlying abnormalities of the spine and spinal cord in many patents with linear scleroderma and a paucity of immunologic abnormalities characteristic of progressive systemic sclerosis. Patients presenting with new onset linear scleroderma should be evaluated for underlying neurologic causes.

Regulation of monocyte/macrophage C2 production and HLA-DR expression by IL-4 (BSF-1) and IFN-gamma.

IL-4 was originally described on the basis of its ability to co-stimulate the proliferation of resting B cells treated with anti-IgM. Recently, this cytokine has been shown to have other effects on mast cells, T cells, B cells, and macrophages. We studied the ability of IL-4 to regulate the production of C2 by human monocytes and monocytic cell lines and compared this with stimulation of HLA-DR expression, another recently described activity of IL-4. Responses to IL-4 were compared to IFN-gamma, a cytokine with both activities. IL-4 up-regulated C2 production by human monocytes and this effect was not inhibited by neutralizing anti-IFN-gamma antibody. IL-4 also stimulated C2 production by HL-60 cells that had been pre-treated with vitamin D3 to induce monocytic differentiation. IL-4 did not stimulate C2 production by U937 cells. IFN-gamma, in contrast to IL-4, stimulates C2 production by all three cell types. Although IL-4 increased C2 production by HL-60 cells we could not detect C2 mRNA by Northern blotting. However, co-stimulation of these cells with IL-4 and low concentrations of IFN-gamma resulted in an additive effect on C2 production and a greater increase in C2 mRNA than was seen with IFN-gamma alone. As reported by others, IL-4-stimulated HLA-DR expression by monocytes. In contrast to our findings regarding C2 production, stimulation of HLA-DR expression was inhibited by neutralizing anti-IFN-gamma mAb and IL-4 did not stimulate HLA-DR expression by U937 or HL-60 cells. IFN-gamma stimulated HLA-DR expression by all three cell types. These results identify IL-4 as an additional cytokine able to directly stimulate C2 production by human monocytes and by a monocytic cell line whereas IL-4 stimulation of HLA-DR expression by monocytes appears to be IFN-gamma dependent.