Precision Medicine Approach for Cardiometabolic Risk Factors in Therapeutic Apheresis.

Lipoprotein apheresis (LA) is currently the most powerful intervention possible to reach a maximal reduction of lipids in patients with familial hypercholesterolemia and lipoprotein(a) hyperlipidemia. Although LA is an invasive method, it has few side effects and the best results in preventing further major cardiovascular events. It has been suggested that the highly significant reduction of cardiovascular complications in patients with severe lipid disorders achieved by LA is mediated not only by the potent reduction of lipid levels but also by the removal of other proinflammatory and proatherogenic factors. Here we performed a comprehensive proteomic analysis of patients on LA treatment using intra-individually a set of differently sized apheresis filters with the INUSpheresis system. This study revealed that proteomic analysis correlates well with routine clinical chemistry in these patients. The method is eminently suited to discover new biomarkers and risk factors for cardiovascular disease in these patients. Different filters achieve reduction and removal of proatherogenic proteins in different quantities. This includes not only apolipoproteins, C-reactive protein, fibrinogen, and plasminogen but also proteins like complement factor B (CFAB), protein AMBP, afamin, and the low affinity immunoglobulin gamma Fc region receptor III-A (FcγRIIIa) among others that have been described as atherosclerosis and metabolic vascular diseases promoting factors. We therefore conclude that future trials should be designed to develop an individualized therapy approach for patients on LA based on their metabolic and vascular risk profile. Furthermore, the power of such cascade filter treatment protocols may improve the prevention of cardiometabolic disease and its complications.

Mutation detection rate and spectrum in familial hypercholesterolaemia patients in the UK pilot cascade project.

Cascade testing using DNA-mutation information is now recommended in the UK for patients with familial hypercholesterolaemia (FH). We compared the detection rate and mutation spectrum in FH patients with a clinical diagnosis of definite (DFH) and possible (PFH) FH. Six hundred and thirty-five probands from six UK centres were tested for 18 low-density lipoprotein receptor gene (LDLR) mutations, APOB p.Arg3527Gln and PCSK9 p.Asp374Tyr using a commercial amplification refractory mutation system (ARMS) kit. Samples with no mutation detected were screened in all exons by single strand conformation polymorphism analysis (SSCP)/denaturing high performance liquid chromatography electrophoresis (dHPLC)/direct-sequencing, followed by multiplex ligation-dependent probe amplification (MLPA) to detect deletions and duplications in LDLR.The detection rate was significantly higher in the 190 DFH patients compared to the 394 PFH patients (56.3% and 28.4%, p > 0.00001). Fifty-one patients had inadequate information to determine PFH/DFH status, and in this group the detection rate was similar to the PFH group (25.5%, p = 0.63 vs PFH). Overall, 232 patients had detected mutations (107 different; 6.9% not previously reported). The ARMS kit detected 100 (44%) and the MLPA kit 11 (4.7%). Twenty-eight (12%) of the patients had the APOB p.Arg3527Gln and four (1.7%) had the PCSK9 p.Asp374Tyr mutation. Of the 296 relatives tested from 100 families, a mutation was identified in 56.1%. In 31 patients of Indian/Asian origin 10 mutations (two previously unreported) were identified. The utility of the ARMS kit was confirmed, but sequencing is still required in a comprehensive diagnostic service for FH. Even in subjects with a low clinical suspicion of FH, and in those of Indian origin, mutation testing has an acceptable detection rate.

Lipoprotein(a): the underutilized risk factor for cardiovascular disease.

Raised lipoprotein(a) [Lp(a)] is an important independent cardiovascular risk factor and predictor of adverse outcomes. Challenges remain with regards to the screening, diagnosis and management of this condition. Although further prospective randomised controlled data is required, there is growing evidence suggesting that lowering Lp(a) may reduce the risk of cardiovascular events and ameliorate symptoms.

Restriction fragment length polymorphisms in the apo B gene in relation to coronary artery disease.

We have determined the frequencies of the alleles at the EcoRI (E), XbaI (X) and PvuII (P) polymorphic restriction sites in the apo B gene in 124 white men with coronary artery disease (CAD) and in 146 white men free from CAD. The frequencies of the E- (restriction site absent) and X- alleles were both significantly higher in normocholesterolaemic men with CAD than in those without CAD, but the frequency of the P+ allele (restriction site present) was similar in the 2 groups. The frequency of the E- allele was significantly higher in CAD men with hypertriglyceridaemia than in normal men without hypertriglyceridaemia. In the normocholesterolaemic men without CAD, the mean serum cholesterol concentration was higher in those with genotype X++ than in those with genotype X--. Mean serum LDL-apo B and LDL-cholesterol concentrations did not differ significantly between men with different XbaI or EcoRI genotypes. Serum apo A-I levels differed significantly between normal men with different XbaI genotypes. Serum HDL-cholesterol levels differed significantly between CAD men with different XbaI genotypes. These results suggest that in white men the E- and X- alleles are in linkage disequilibrium with a nearby allele that is causally related to CAD. It is also possible that the amino acid substitution at position 4154 in apo B, brought about by the nucleotide change responsible for the EcoRI polymorphism, has a direct effect on the atherogenicity of LDL.

Apolipoprotein A-I gene polymorphisms: frequency in patients with coronary artery disease and healthy controls and association with serum apo A-I and HDL-cholesterol concentration.

We have investigated the association between serum high density lipoprotein-cholesterol (HDL-C) and apo A-I concentration and the PstI and XmnI restriction fragment length polymorphisms of the apolipoprotein AI-CIII-AIV multigene complex. Two groups of subjects were examined. The first comprised 174 unrelated male patients under 60 years of age with angiographic evidence of coronary artery disease (CAD). Of this group 34 were non-North European. The second group consisted of 104 unrelated healthy male North European subjects aged under 60 and free from demonstrable CAD, who attended a health screening clinic in London. For the PstI polymorphism, the frequency of the rarer P2 allele was 0.12 in both the North European and non-North European patients and this was higher than in the control group (P2 frequency 0.06, P less than 0.05). Healthy individuals with the genotype P1P2 had higher levels of apo A-I but similar levels of HDL-C compared to those with the genotype P1P1. However, CAD patients with the genotype P1P2 had lower serum levels of apo A-I and significantly lower serum levels of HDL-C compared to those with the genotype P1P1 (0.85 mmol/l vs. 1.0 mmol/l, P less than 0.05). The allele frequencies of the XmnI polymorphisms were not significantly different in the control group and the group of North European patients, although within the sample of non-North European patients, the frequency of the X2 allele was significantly higher than that found in the North European controls (0.26 vs. 0.09). Patients with the genotype X1X2 had a higher mean serum concentration of HDL-C and apo A-I compared with patients with the genotype X1X1 (1.14 and 0.93 mmol/l for HDL-C, P less than 0.05; 147 and 123 mg/dl for apo A-I, P less than 0.05). Associations between HDL-C and apo A-I levels and PstI and XmnI genotype were similar in patients taking and not taking beta-blockers. The data show that genetic variation in the apo AI-CIII-AIV gene cluster is associated with coronary artery disease although only weakly, and suggest that the mechanism of this association may operate through an effect in determining the serum concentration of apo A-I and HDL-cholesterol.

Determinants of transplant-related coronary calcium detected by ultrafast computed tomography scanning.

Coronary calcium detected by ultrafast computed tomography (CT) has been shown to be a marker of coronary artery disease in heart transplant recipients. The objective of this study was to examine the possible determinants of coronary calcium after heart transplantation. Over a 15-month period, 102 consecutive cardiac transplant recipients (mean age 53 years, 88 men) underwent ultrafast CT scanning of the heart, in addition to coronary angiography, to determine coronary calcium score on their annual follow-up (a median of 4.6 years [range 63 days to 9.1 years] after transplant). The following data were also recorded: the recipient's sex and date of birth, date of transplantation, date of ultrafast computed tomography and coronary angiography; recipient pretransplant diagnosis, history of diabetes mellitus and systemic hypertension, fasting lipid profile, immunosuppression, number of rejection episodes, and donor organ ischemic time. Forty six patients (45.1%) had total calcium scores >0 and 41 (40.2%) had at least 1 major coronary with angiographic narrowing >24%. On univariate analysis, coronary calcium was significantly associated with dyslipoproteinemia, total cholesterol was >6.0 mmol/L (240 mg/dl), triglycerides were >3.0 mmol/L (265 mg/dl), and lipoprotein(a) >30 mg/ dl; > or =25% angiographic disease was significantly associated with coronary calcium and dyslipoproteinemia. Logistic regression revealed that dyslipoproteinemia, systemic hypertension, and donor ischemic time were significant predictors of coronary calcium in transplanted hearts. We conclude that the prevalence of coronary calcium in heart transplant recipients is high and is related to recipient dyslipoproteinemia, systemic hypertension, and donor organ ischemic time.

Maxepa versus bezafibrate in hyperlipidemic cardiac transplant recipients.

Accelerated coronary artery disease is the most serious obstacle to long-term survival in cardiac transplant recipients. Lipid abnormalities are found frequently in these patients, and there is growing evidence that even minimally increased levels of cholesterol and triglycerides contribute to the development of accelerated coronary artery disease. However, the optimal lipid-lowering therapy after cardiac transplantation has not been defined. In an open, randomized study, the efficacy and safety of bezafibrate (400 mg/day) and fish oil (Maxepa) (10 g/day) for 3 months were compared in 87 cardiac transplant recipients with serum total cholesterol > 6.5 or triglycerides > 2.8 mmol/liter, or both. After 1 month, bezafibrate reduced total cholesterol by 13%, low-density lipoprotein cholesterol by 20% and apolipoprotein B by 13%. It also increased apolipoprotein A1 and high-density lipoprotein cholesterol by 12 and 20%, respectively, and significantly reduced fibrinogen at 3 months. Maxepa had no significant effect on these variables, but was as effective as bezafibrate in reducing triglycerides (36 and 31%, respectively). Both drugs increased lipoprotein (a) to a similar extent, and bezafibrate significantly increased serum creatinine. These results suggest that bezafibrate has better lipid-, apolipoprotein- and hemostatic modifying properties than does Maxepa, but its potentially adverse effect on renal function needs further investigation.

Antiendothelial antibodies after heart transplantation: the accelerating factor in transplant-associated coronary artery disease?

Although the precise cause of transplant-associated coronary artery disease is unknown, immune mechanisms have been implicated. Using the techniques of SDS-PAGe and Western immunoblotting, we have previously shown that a strong positive correlation exists between the development of coronary artery disease and the presence of antiendothelial antibodies reactive with a doublet of polypeptides of approximately 60 and 62 kDa. We have now extended this study to investigate the temporal pattern of antiendothelial antibody formation after transplantation and its association with cellular rejection episodes. The original study used patients in whom coronary artery disease had developed early after transplantation, that is at 1 or 2 years. Here we investigate whether antiendothelial antibodies are also made in patients in whom the disease does not develop until 5 to 10 years after heart transplantation and whether the antibodies are found in patients with severe nontransplant atherosclerosis. We confirm the 60 to 62 kDa antigens are membrane bound, and recalculation of their molecular mass makes the doublet 56 and 57.5 kDa. The results show that antibodies specific for the doublet of endothelial antigens are rarely produced by patients other than those in whom rapidly progressing coronary artery disease develops early after transplantation. The antibodies are unrelated to cellular rejection episodes. We believe their production may be an accelerating factor for the rapid development of transplant-associated coronary artery disease.

Combined bezafibrate and simvastatin treatment for mixed hyperlipidaemia.

The safety and efficacy of combined bezafibrate-simvastatin therapy was evaluated in 49 patients with diet-resistant mixed hyperlipidaemia (type IIb). After a two-month placebo phase, patients were randomized to receive either Bezafibrate Slow Release (SR) 400 mg mane or simvastatin 20 mg nocte followed by three months combination therapy. Total cholesterol, triglycerides and high-density lipoprotein (HDL) cholesterol were measured at monthly intervals. Apolipoproteins (apo) A1 and B, lipoprotein (a) [Lp(a)] and fibrinogen were measured before and after each treatment. Simvastatin was more effective than Bezafibrate SR in reducing total cholesterol (2.0 vs. 1.1 mmol/l, p = 0.003) and lowering LDL cholesterol (1.7 vs. 0.4 mmol/l, p = 0.0001) whereas Bezafibrate SR was more effective in reducing triglycerides (by 41% vs. 17%, p = 0.001) and fibrinogen (by 23% vs. 3%, p = 0.004). Compared with simvastatin monotherapy, combined drug therapy induced further reductions in triglycerides (by 26%, p = 0.0003) and apoB (by 11 mg/dl, p = 0.03) and an increase in apoA1 (by 21 mg/dl, p = 0.0008). Symptomatic and biochemical adverse events did not occur more frequently on combined drug therapy than on monotherapy. The combination of bezafibrate and simvastatin was more effective in controlling mixed hyperlipidaemia than either drug alone and did not provoke more adverse events.

Coronary endothelial function in cardiac transplant recipients with accelerated coronary disease.

BACKGROUND: Previous studies with the endothelium-dependent vasodilator substance P have shown a preserved vasodilator response in cardiac transplant recipients with angiographically normal coronary arteries. Although endothelial dysfunction is known to occur in cardiac transplant recipients with accelerated coronary disease, the degree to which the endothelium is affected is not known precisely. The aim of the present study was to examine endothelial function in accelerated coronary disease following cardiac transplantation. METHODS: Thirteen cardiac transplant recipients with epicardial coronary disease underwent substance P infusion. The response to incremental doses of substance P was measured in smooth segments of affected coronary arteries. Substance P was infused over 2 min with a starting dose of 1.4 pmol/min and a maximum of 22.4 pmol/min, reached by doubling the dose in steps, followed by an infusion of 2 mg isosorbide dinitrate over 2 min. RESULTS: Substance P caused less vasodilation at lower concentrations, with a significantly higher dose required to achieve half maximal dilation compared with cardiac transplant recipients with no coronary disease. The mean maximal dilatation achieved with substance P was 22.98 +/- 4.62% compared to 21.95 +/- 4.9% with isosorbide dinitrate; the latter value was not significantly different from the maximal dilation achieved in cardiac transplant recipients without coronary disease. CONCLUSIONS: In cardiac transplant recipients with accelerated coronary disease the functional vasodilatory ability of the coronary endothelium is impaired in segments of apparently unaffected epicardial arteries, which may lead to an increase in the resting vasoconstrictor tone and have important functional and therapeutic implications.

Magnetic resonance coronary angiography in heart transplant recipients.

BACKGROUND: Magnetic resonance angiography (MRA) using segmented k-space fast low-angle shot imaging has recently been used to demonstrate the proximal coronary arteries in healthy subjects and in patients with coronary artery disease. We assessed the sensitivity and specificity of coronary MRA in heart transplant recipients and investigated the feasibility of coronary MRA in patients with metallic sutures and clips in the chest. MATERIALS AND METHODS: Sixteen cardiac transplant patients aged 57.2 +/- 7.9 years (mean +/- SD) were recruited. Forty-eight arterial segments were evaluated, including the left main artery (LMA), left anterior descending artery (LADA) and right coronary artery (RCA). We excluded the left circumflex artery which could not be imaged accurately. The average time between heart transplant operation and MRA was 6 years, whereas that between MRA and X-ray angiography was 4 months. The coronary MRA was interpreted by two experienced investigators who were blinded to the coronary X-ray angiography results. Similarly, the coronary X-ray angiography results were interpreted by two experienced investigators blinded to the MRA results. The coronary arterial segments were classified by MRA as being normal or as having an amount of disease that was significant (> 50% lesion) or insignificant (< 50% lesion). RESULTS: There were 28 true-negative, five true-positive, four false-negative and six false-positive results. Of the 28 true-negative cases, 13 were in the LMA, six in the LADA and nine in the RCA. There was one false-positive LMA, two false-positive LADA and three false-positive RCA stenoses. There were four false-negative results in the LADA and one in the RCA. Clips precluded evaluation in one LMA, one LADA and one RCA. One LMA and one LADA were not evaluated as a result of poor images. One false-positive RCA stenosis was caused by a metallic clip. Three of the false-negative LADA stenoses had lesions in the distal third of the artery. The sensitivity, specificity, negative and positive predictive values were generally poor for the left coronary artery. The best results were for the RCA (sensitivity 100%, specificity 75%, positive predictive value 50% and negative predictive value 100%). The specificity in the left coronary arteries (LMA and LADA) was 86%, but the other indicators were all poorer. For the RCA, LMA and LADA combined, the overall sensitivity was 56%, specificity 82%, predictive accuracy 45% and negative predictive value 88%. In three patients, < 50% RCA lesions were seen in the MRA data, which were all confirmed by angiography. No < 50% lesions were seen in the LMA or in the LADA by MRA or by X-ray angiography. CONCLUSION: Coronary MRA using the segmented fast low-angle shot technique is feasible in heart transplant recipients but the sensitivity and specificity of this method are limited. Further developments in coil design, rapid imaging techniques and respiratory monitoring methods are necessary to improve the accuracy of coronary MRA.

Relationship of immunosuppression and serum lipids to the development of coronary arterial disease in the transplanted heart.

Coronary arterial disease in the cardiac allograft has emerged as the most serious long term complication of cardiac transplantation. The influence of patient-related and other potential risk factors on the development of coronary arterial disease at 1 year subsequent to cardiac transplantation was examined in 207 recipients. The mean age of donors in patients with coronary arterial disease was 28.5 +/- 9.5 years, compared to 22.6 +/- 7.9 years in patients without coronary arterial disease (P less than 0.01). Eight of the 35 patients who received immunosuppression by means of prednisone and azathioprine developed coronary arterial disease compared to 5 of the 172 patients who were treated with cyclosporin and azathioprine without routine oral prednisone (P less than 0.01). The relationship of levels of serum lipids to the subsequent development of coronary arterial disease was investigated in 95 patients with angiographically normal coronary arteries one year after cardiac transplantation. The cumulative probability of coronary arterial disease in those with total cholesterol greater than 5.8 mmol/l was 9.3% at 2 years (n = 40), 24.4% at 4 years (n = 21) and 45% at 4 years (n = 9) compared with 4.3% at 2 years (n = 45), 7.4% at 3 years (n = 32) and 14% at 4 years (n = 14) in those with a total cholesterol less than 5.8 mmol/l (P less than 0.05). Similarly, the incidence of coronary arterial disease was increased in patients with serum triglyceride greater than 1.4 mmol/l (P less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Efficacy and safety of fluvastatin therapy for hypercholesterolemia after heart transplantation: results of a randomised double blind placebo controlled study.

BACKGROUND: Hypercholesterolemia is frequent after heart transplantation. Statins can reduce cholesterol levels but their use in heart transplant patients is complicated by pharmacokinetic interactions with cyclosporin and the risk of serious adverse effects including rhabdomyolysis. Fluvastatin has been used safely to treat hypercholesterolemia in renal transplant patients but there are few data relating to its use after heart transplantation. Therefore, we conducted a randomised blinded placebo controlled trial. METHODS AND RESULTS: Seventy-nine patients, 3 months to 12 years after heart transplantation with a low density lipoprotein (LDL) cholesterol between 3.5 and 8.0 mmol/l were randomly assigned, in a 2:1 ratio, to receive either fluvastatin 40 mg od (n=52) or matching placebo (n=27). Changes in total cholesterol (TC) in the fluvastatin and placebo groups were -17.0% and +4.5%, respectively, (p<0.001); the corresponding changes in LDL were -20.5% and +4.8% (P<0.001) and in triglycerides -14.5% and +7.1% (p=0.012) at the end of the 1-year study period. Withdrawals were more frequent in the fluvastatin group (23% vs. 11% p=0.24). Two deaths occurred during the study (the rate expected from International Society of Heart Lung Transplantation registry) and appeared to be unrelated to the study medication. There were no episodes of rhabdomyolysis or other serious drug-related side effects. CONCLUSIONS: Fluvastatin (40 mg/day) was both an effective and a safe treatment for hypercholesterolemia in patients who had undergone heart transplantation more than 3 months previously.

Low-dose simvastatin for the treatment of hypercholesterolaemia in recipients of cardiac transplantation.

There is increasing evidence that hypercholesterolaemia is an important contributor to the development of accelerated coronary arterial disease in the cardiac allograft. The optimal drug therapy of hypercholesterolaemia in recipients after cardiac transplantation, however, has not been defined. Simvastatin (an inhibitor of hydroxy-methyl glutaryl-coenzyme A reductase), at a dose of 10 mg/day, was administered to 12 recipients with serum total cholesterol greater than or equal to 7.8 mmol/l and serum triglyceride less than or equal to 4.5 mmol/l refractory to dietary measures during a follow-up period of 1-5 years after cardiac transplantation. All patients received maintenance doses of cyclosporin A and, in some instances, azathioprine and prednisolone. After 2 months treatment with simvastatin, serum total cholesterol was significantly reduced from 8.8 +/- 0.3 mmol/l (mean +/- SEM) to 5.5 +/- 0.5 mmol/l, P less than 0.001, low density cholesterol from 6.6 +/- 0.4 to 3.8 +/- 0.3 mmol/l, P less than 0.001 and triglycerides from 2.4 +/- 0.2 mmol/l to 1.8 +/- 0.2 mmol/l, P less than 0.005. These changes were maintained after a period of treatment of 8 months. Serum high density cholesterol, hepatic transaminase levels, serum creatinine, creatine kinase and cyclosporin A blood levels were not altered by treatment with simvastatin. It is concluded that, in this study group, low-dose simvastatin appears to be well tolerated and has favourable lipid modifying properties.

A randomized pilot trial of low-dose combination lipid-lowering therapy following coronary artery bypass grafting.

Vein graft atherosclerosis is a common and serious complication of coronary artery bypass grafting (CABG). There is mounting evidence that lipoprotein abnormalities play an equally important role in the development of lesions in saphenous vein grafts after CABG as in native coronary vessel disease. The potential benefit of low-dose lipid lowering combination therapy in these patients has not been investigated. In a randomized, double-blind, placebo-controlled study, we compared the efficacy and safety of a low-dose combination of colestipol 10 g and simvastatin 10 mg/day (CS) to colestipol 10 mg and bezafibrate 400 mg/day (CB) for 2 months in 33 patients with serum total cholesterol > 6.5 mmol/l and triglyceride < 4.5 mmol/l who had undergone CABG for severe coronary artery disease. In the CS group, total cholesterol decreased by 29% and low-density lipoprotein (LDL) cholesterol by 42%; similarly, CB reduced total cholesterol by 17%, LDL cholesterol by 23%, triglyceride by 19%, and increased high-density lipoprotein (HDL) cholesterol by 14%. Lipoprotein (a) and hemostatic factors were unaffected by either therapy in this study. Both combination therapies were well tolerated with no significant clinical or biochemical side effects. We conclude that low-dose combinations of colestipol and simvastatin or colestipol and bezafibrate are effective and well tolerated in the management of moderate hyperlipidemia in patients who had undergone CABG.

Efficacy criteria and cholesterol targets for LDL apheresis.

Low density lipoprotein (LDL) apheresis is now accepted as the treatment of choice for patients with homozygous familial hypercholesterolaemia and for heterozygotes with cardiovascular disease refractory to lipid-lowering drug therapy. However, a paucity of evidence has meant that detailed guidance on the extent of cholesterol reduction required to prevent the onset or progression of cardiovascular disease in these high risk patients is lacking. This review defines criteria for expressing the efficacy of apheresis, proposes target levels of total and LDL cholesterol for homozygotes and heterozygotes based on recent follow-up studies and suggests a scheme for monitoring cardiovascular disease in these patients. Establishing a uniform approach to data collection would facilitate the setting up of national or multi-national registers and might eventually provide the information needed to formulate evidence-based guidelines for LDL apheresis.