In vivo hypertensive arterial wall uptake of radiolabeled liposomes.

Using five sham-operated and seven aortic coarctation-induced hypertensive New Zealand White rabbits intravenously injected with neutral small unilamellar vesicles loaded with [111In]nitrilotriacetic acid, we demonstrated in vivo that the normal aortic arterial wall participates in liposome uptake and that this uptake is increased in the hypertensive aortic wall by approximately threefold (p less than or equal to 0.0001). Among the three regions examined, aortic arch, thoracic aorta, and lower abdominal aorta, the difference in uptake between the normotensive and hypertensive arterial walls was significantly different, p less than or equal to 0.05, p less than or equal to 0.0001, and p less than 0.05, respectively. The uptake by the different regions of the hypertensive arterial wall is consistent with the pathological changes present in these areas. Furthermore, the extent of liposome uptake by the aortic wall is strongly correlated with the height of the blood pressure (r = 0.85, p = 0.001, n = 11). We conclude that neutral small unilamellar liposomes can be used to carry agents into the arterial wall in vivo in the study of hypertensive vascular disease and could be especially useful for the delivery of pharmacologically or biologically active agents that would otherwise be inactivated within the circulation or are impermeable to the arterial wall.

Relationship of arterial wall uptake of radiolabeled liposomes to the presence of monocyte/macrophage cells in the hypertensive and atherosclerotic arterial wall.

In vivo radiolabeled liposome uptake in 5 sham-operated, 7 coarctation-induced hypertensive, and 8 atherosclerotic arterial walls from New Zealand White rabbits was compared to determine the mechanism of arterial wall uptake of liposomes. Uptake between the three groups was significantly different (P less than 0.001) with a 3-fold difference in uptake between the sham-operated and hypertensive groups and the hypertensive and atherosclerotic groups. Liposome uptake was significantly higher in the atherosclerotic group of animals (P less than 0.05). Avidin-biotin immunoperoxidase staining for monocyte/macrophage cells revealed that liposome uptake increased concomitantly with arterial wall monocyte/macrophage cellular invasion and that liposome localization, determined by autoradiography, paralleled the monocyte/macrophage cellular distribution in both hypertensive and atherosclerotic arterial walls. This study provides the first direct evidence that liposomes can escape from the circulation and enter the diseased arterial wall. Furthermore, it suggests that one possible mechanism of arterial wall uptake of liposomes is via the monocyte/macrophage cell which avidly and preferentially engulfs liposomes and then passively carries them into the arterial wall during hypertensive and atherosclerotic lesion development. Liposomes could potentially be used to carry agents into the arterial wall in the study of arterial wall lesion development.

Prediction of angiographic change in native human coronary arteries and aortocoronary bypass grafts. Lipid and nonlipid factors.

A within-group risk factor analysis was conducted to predict angiographic change in the Cholesterol Lowering Atherosclerosis Study, a randomized, placebo-controlled trial of colestipol plus niacin therapy in men with previous coronary bypass surgery. Global angiographic change, including both native coronary arteries and bypass grafts after 2 treatment years, was the end point. Risk factors included on-trial clinical measures, plasma lipids, lipoproteins, and apolipoproteins. Univariate analysis indicated that risk factors previously observed by others in epidemiologic investigation of ischemic heart disease--total cholesterol, LDL cholesterol, non-HDL cholesterol, triglycerides, apolipoprotein B, and diastolic blood pressure--had significant effects in the placebo-treated group. Univariate analysis indicated significant effects of apolipoprotein C-III in drug- and placebo-treated groups. Multivariate analysis indicated the predominant risk factor predicting the probability of global coronary progression was non-HDL cholesterol in placebo-treated subjects and the content of apolipoprotein C-III in high density lipoproteins of drug-treated subjects. Both drug- and placebo-treated group findings point to an important role for triglyceride-rich lipoproteins in progression and regression of human atherosclerosis.

Combined therapy of niacin, colestipol, and fat-controlled diet in men with coronary bypass. Effect on blood lipids and apolipoproteins.

The effects of colestipol (30 grams/day), niacin (7.3 grams/day), and diet on blood lipids and apolipoproteins after one year of therapy are reported. Men selected on the basis of previous coronary artery bypass surgery were randomly assigned to drug or control treatments in an angiographic study of atherosclerosis progression and regression. In 14 men, drugs and diet produced the following changes: Baseline total cholesterol 245 mg/dl, triglyceride 189 mg/dl, and LDL cholesterol 164 mg/dl were decreased by 73 mg/dl (29%), 83 mg/dl (41%) and 69 mg/dl (40%) respectively. Baseline HDL cholesterol, 44 mg/dl was increased 13 mg/dl (33%). Baseline apolipoprotein B, 124 mg/dl and apolipoprotein C-III (heparin precipitate) 5.6 mg/dl were decreased 40 mg/dl (31%) and 2.4 mg/dl (41%) respectively. All these changes are significant, p less than 0.01. Apolipoprotein A-I and apolipoprotein C-III (heparin supernate) were not significantly changed. In the controls, placebo and diet produced no significant decrease in blood lipid or lipoproteins, with the exception that baseline apolipoprotein B, 111 mg/dl increased 18 mg/dl (12%), p less than 0.05.

Beneficial effects of combined colestipol-niacin therapy on coronary atherosclerosis and coronary venous bypass grafts.

The Cholesterol-Lowering Atherosclerosis Study (CLAS) was a randomized, placebo-controlled, angiographic trial testing combined colestipol hydrochloride and niacin therapy in 162 nonsmoking men aged 40 to 59 years with previous coronary bypass surgery. During two years of treatment there was a 26% reduction in total plasma cholesterol, a 43% reduction in low-density lipoprotein cholesterol, plus a simultaneous 37% elevation of high-density lipoprotein cholesterol. This resulted in a significant reduction in the average number of lesions per subject that progressed (P less than .03) and the percentage of subjects with new atheroma formation (P less than .03) in native coronary arteries. Also, the percentage of subjects with new lesions (P less than .04) or any adverse change in bypass grafts (P less than .03) was significantly reduced. Deterioration in overall coronary status was significantly less in drug-treated subjects than placebo-treated subjects (P less than .001). Atherosclerosis regression, as indicated by perceptible improvement in overall coronary status, occurred in 16.2% of colestipol-niacin treated vs 2.4% placebo treated (P = .002).

The Cholesterol Lowering Atherosclerosis Study (CLAS): design, methods, and baseline results.

The Cholesterol Lowering Atherosclerosis Study (CLAS) is a prospective, placebo-controlled, angiographic trial designed to test the hypothesis that aggressive lowering of LDL cholesterol with concomitant increase in HDL cholesterol will reverse or retard the atherosclerotic process. Specifically, CLAS was designed to determine whether combined therapy with colestipol plus niacin will produce clinically significant change in coronary, carotid, and femoral artery atherosclerosis and coronary bypass graft lesions. To this purpose, 188 subjects were randomized to diet plus drug or diet plus placebo. We report on methodological aspects of planning and evaluating this study, including the choice of the study population, procedures for recruitment, the experimental design including sample size considerations, methods for evaluating outcome, and methods for evaluating compliance to treatment. Comparison of baseline data indicated no significant differences between groups at the time of randomization. Subjects were predominantly male, Caucasian, 54 years of age, 20% above ideal weight, with normal blood pressure. The average age at bypass was 50 years. The average lipids were cholesterol (243 mg/dL), HDL (45 mg/dL), and LDL (168 mg/dL). Finally, the distribution of baseline coronary stenosis was equivalent between the two groups (average number of lesions per subject = 10.6).

Beneficial effects of colestipol-niacin on coronary atherosclerosis. A 4-year follow-up.

The Cholesterol Lowering Atherosclerosis Study (CLAS) was a randomized, placebo-controlled, angiographic trial testing combined colestipol-niacin therapy in 162 subjects. Two-year results (CLAS-I) showed decreased atherosclerosis progression and increased regression. We now describe a subgroup of 103 subjects treated for 4 years (CLAS-II). Changes in blood lipid, lipoprotein-cholesterol, and apolipoprotein levels were maintained, and at 4 years significantly more drug-treated subjects demonstrated nonprogression (52% drug- vs 15% placebo-treated) and regression (18% drug- vs 6% placebo-treated) in native coronary artery lesions. Significantly fewer drug-treated subjects developed new lesions in native coronary arteries (14% drug- vs 40% placebo-treated) and bypass grafts (16% drug- vs 38% placebo-treated). These results confirm CLAS-I findings and indicate that regression can continue for 4 years. They reaffirm the need for early initiation of vigorous long-term lipid lowering therapy in coronary bypass subjects.

Beneficial effects of colestipol-niacin therapy on the common carotid artery. Two- and four-year reduction of intima-media thickness measured by ultrasound.

BACKGROUND: Controlled clinical trials have reported treatment effects evaluated with serial imaging in coronary and femoral but not cervical arteries. The Cholesterol Lowering Atherosclerosis Study, a coronary, cervical, and femoral angiographic trial of colestipol plus niacin, included a pilot study of standardized carotid ultrasound imaging. METHODS AND RESULTS: Seventy-eight subjects had ultrasound studies at baseline, 2, and 4 years. Twenty-four drug and 22 placebo subjects had carotid ultrasound images at baseline, 2, and 4 years with matching cervical angiograms. Computer image processing was applied to ultrasound images of common carotid (far wall) and cervical angiograms. Computer operators were blind to treatment group. Carotid ultrasound measurements were tested for treatment effects and compared with measurements of atherosclerosis in coronary and cervical angiograms. Drug subjects showed significant progressive reduction in carotid thickness at 2 (P = .0001) and 4 years (P = .0001); placebo subjects significantly increased wall thickness at 2 and 4 years. Reduced levels of apolipoprotein B and increased levels of high density lipoprotein cholesterol and apolipoprotein C-III were significant predictors of carotid wall thinning. Ultrasound-measured carotid intima-media thickness was correlated at baseline with visually read coronary angiographic stenosis and at 2 years with a robust computer measurement of mild carotid atherosclerosis. CONCLUSIONS: Common carotid intima-media thickening can be reduced by colestipol-niacin treatment. Two-year image-processed carotid ultrasound trials can provide adequate power with 50 subjects per group to test for this treatment effect.

Evaluation of human panelists in assessing coronary atherosclerosis.

The Cholesterol Lowering Atherosclerosis Study, a randomized, angiographic clinical trial, has demonstrated the beneficial effect of niacin/colestipol therapy on coronary and femoral atherosclerosis. The primary outcome was a panel-determined consensus score evaluating global coronary changes determined angiographically at 2 years. This article presents an evaluation of interreader agreement in independently assessing the status of native coronary arteries and overall coronary condition. Parameters include 1) identification of the presence of lesions and lesion changes; 2) estimation of lesion severity (percent stenosis) and amount of change in lesion severity; and 3) global assessment of change in coronary status. Readers independently agreed on 1) presence of lesions (82%) and change in lesions (51%); 2) percent stenosis +/- 10% (76%) and change in stenosis +/- 10% (81%); and 3) global assessment of change in coronary status within one step (96%). Results of these analyses may be useful in effectively designing angiographic trials that use a panel of human evaluators as well as computerized methods for angiographic interpretation.

Effects of colestipol-niacin therapy on human femoral atherosclerosis.

The 2-year therapy effect on femoral atherosclerosis was evaluated in the Cholesterol Lowering Atherosclerosis Study (CLAS), a randomized, placebo-plus-diet-controlled angiographic trial of colestipol-niacin therapy plus diet in men with previous coronary bypass surgery. Different diet compositions were prescribed to enhance the differential in blood cholesterol responses between the two groups. The annual rate of change in computer-estimated atherosclerosis (CEA), a measure of lumen abnormality, was evaluated between treatment groups. A significant per-segment therapy effect was found in segments with moderately severe atherosclerosis (p less than 0.04) and in proximal segments (p less than 0.02). When segmental CEA measures were combined into a per-patient score using an adaptation of the National Heart, Lung, and Blood Institute scoring procedure, a significant therapy effect was observed (p less than 0.02). Total variance of the annual change rate in CEA was as predicted from pilot studies, but measurement variation was larger. The therapy effect observed in femoral arteries, although significant, was less marked than the strong and consistent benefit previously reported for both native coronary arteries and aortocoronary bypass grafts.

Progression of coronary artery disease predicts clinical coronary events. Long-term follow-up from the Cholesterol Lowering Atherosclerosis Study.

BACKGROUND: Progression of coronary artery disease is assumed to be a surrogate end point for clinical coronary events. Because no single method or measure for a coronary angiographic end point is uniformly accepted as optimal, the utility and validity of surrogate end points for predicting clinical coronary events remain unsettled. METHODS AND RESULTS: The Cholesterol Lowering Atherosclerosis Study randomized 162 nonsmoking, 40- to 59-year-old men with previous coronary artery bypass graft surgery to colestipol/niacin plus diet or placebo plus diet. Atherosclerosis change on 2-year coronary angiograms was evaluated by a consensus panel and by quantitative coronary angiography (average per-subject change in percent diameter stenosis [%S] and minimum lumen diameter [MLD). With all three end points, the benefit of colestipol/niacin treatment on coronary artery atherosclerosis has been reported. Annual follow-up for an average of 7 years (range, 6.3 months to 10 years) has been carried out on all subjects who completed the 2-year angiogram. Clinical coronary events (need for revascularization, nonfatal acute myocardial infarction, and coronary death) have been documented. Risk of clinical coronary events was positively related to coronary lesion progression for all three surrogate end points (P<.05). New lesion formation in bypass grafts (P=.02) and progression of mild/moderate lesions ( < 50%S) were predictive of clinical coronary events (P<.01). Change in MLD contributed significantly to the prediction of clinical coronary events beyond a model with %S alone (P<.05). CONCLUSIONS: In this population of nonsmoking men with previous bypass surgery, both the consensus panel- and quantitative coronary angiography-based end points of coronary artery disease progression predict clinical coronary events. Subjects who demonstrate greater coronary artery lesion progression have an increased risk of future clinical coronary events. Design of shorter, smaller trials of antiatherosclerotic agents is justified.

Alterations in serum thyroid hormonal indices with colestipol-niacin therapy.

A serial blood-lipid-lowering study at the University of Southern California yielded unexpected findings on routine thyroid function monitoring. After 1 year of combined colestipol and niacin therapy, patients had reduced total serum thyroxine (T4) levels and increased triiodothyronine uptake ratios, an indicator of apparent decreases in thyroxine-binding globulin levels. Calculation of the free T4 index partially but not completely corrected for the apparent decrease in thyroxine-binding globulin, as determined by a relatively small decrease in the free T4 index compared with a large decrease in T4. Sequential sampling, using three separate methods, showed reduced thyroxine-binding globulin levels. The mechanism for these changes is unknown, but the fact that these patients were essentially euthyroid needs emphasis because the use of combined colestipol and niacin therapy is becoming more widespread.

Comparison of computer- and human-derived coronary angiographic end-point measures for controlled therapy trials.

The Cholesterol Lowering Atherosclerosis Study, a randomized angiographic clinical trial, demonstrated the beneficial effect of niacin/colestipol plus diet therapy on coronary atherosclerosis. Outcome was determined by panel-based estimates (viewed in both still and cine modes) of percent stenosis severity and change in native artery and bypass graft lesions. Computer-based quantitative coronary angiography (QCA) was also used to measure lesion and bypass graft stenosis severity and change in individual frames closely matched in orientation, opacification, and cardiac phase. Both methods jointly evaluated 350 nonoccluded lesions. The correlation between QCA and panel estimates of lesion size was 0.70 (p less than 0.0001) and for change in lesion size was 0.28 (p = 0.002). Agreement between the two methods in classifying lesion changes (i.e., regression, unchanged, or progression) occurred for 60% (210 of 350) of the lesions kappa +/- SEM = 0.20 +/- 0.05, p less than 0.001). The panel identified 442 nonoccluded lesions for which QCA stenosis measurements could not be obtained. Lesions not measurable by QCA included those with stenosis greater than 85% that could not be reliably edge tracked, segments with diffuse or ecstatic disease that had no reliable reference diameter, and segments for which matched frames could not be located. Seventy-nine lesions, the majority between 21% and 40% stenosis, were identified and measured by QCA but were not identified by the panel. This comparison study demonstrates the need to consider available angiographic measurement methods in relation to the goals of their use.

Evaluation of colestipol/niacin therapy with computer-derived coronary end point measures. A comparison of different measures of treatment effect.

BACKGROUND: The Cholesterol Lowering Atherosclerosis Study has demonstrated beneficial effect of colestipol/niacin on coronary atherosclerosis using a panel-determined global coronary change score. We now report treatment group comparisons using quantitative coronary angiographic (QCA) measures from all processable segments in 85 of 162 randomly selected baseline/2-year film pairs. METHODS AND RESULTS: Treatment benefit was established for percent stenosis for either continuous or categorical analyses with regression established regardless of the per-patient scoring procedure. In addition, treatment benefit favoring regression was established in some cases for roughness and for percent involvement, a longitudinal estimate of the percent of coronary surface involved by raised lesions. Benefit on minimum diameter was directly related to whether the segment was proximal to a graft insertion and hemodynamically related to the bypass graft. QCA correlates of panel-determined progression were increases in percent stenosis and numbers of occluded lesions in native arteries and the number of progressing lesions in bypass grafts. CONCLUSIONS: These results demonstrate that a variety of computer measures can be used as end points in coronary angiographic therapy trials, but change in percent stenosis correlates best with visual panel assessments and best reflects the treatment benefit; when treatment effect sizes are moderate to large, the required sample size of coronary angiographic trials can be reduced when QCA is used.

One-year reduction and longitudinal analysis of carotid intima-media thickness associated with colestipol/niacin therapy.

BACKGROUND AND PURPOSE: The Cholesterol Lowering Atherosclerosis Study has reported significant reduction of coronary artery disease and of carotid arterial intima-media thickness (IMT) at 2 and 4 years with colestipol/niacin therapy. We now report on treatment effects on carotid IMT at 6 months and 1 year. METHODS: One hundred eighty-eight nonsmoking men, aged 40 to 59 years, with prior coronary artery bypass graft surgery were randomized to colestipol/niacin plus diet therapy or placebo plus diet therapy. Computerized image processing of carotid ultrasound films was used to measure IMT in the right common carotid artery. Treatment group comparisons were made at 6 months and 1 year (46 and 33 subjects, respectively, with baseline and 6-month or 1-year ultrasound measures). The time course of the treatment effect on carotid IMT was estimated using the complete sample of 78 subjects with baseline and on-trial data. RESULTS: No significant treatment group differences on carotid IMT were found at 6 months. At 1 year, the treated group showed significant reduction of carotid IMT (P = .01 between groups). The placebo group showed continuing progression of IMT during the 4-year study period (estimated progression rate, 0.018 mm/y). The treated group showed reduction of IMT during the first 3 years and a plateau during the remainder of the study. CONCLUSIONS: Reduction of carotid IMT was found with aggressive lipid-lowering therapy. Ultrasound measures of IMT offer a noninvasive and precise measure of early carotid atherosclerosis that will decrease sample size requirements, potentially decrease dropout rates, and widen the study population of antiatherosclerotic clinical trials.