Effects of sirolimus on plasma lipids, lipoprotein levels, and fatty acid metabolism in renal transplant patients
2002; Elsevier BV; Volume: 43; Issue: 8 Linguagem: Inglês
10.1194/jlr.m100392-jlr200
ISSN1539-7262
AutoresJoel D. Morrisett, Ghada Abdel‐Fattah, Ron C. Hoogeveen, Eddie Mitchell, Christie M. Ballantyne, Henry J. Pownall, Antone R. Opekun, Jonathon S. Jaffe, Suzanne Oppermann, Barry D. Kahan,
Tópico(s)Liver Disease and Transplantation
ResumoSirolimus (Rapammune®, rapamycin, RAPA) is a potent immunosuppressive drug that reduces renal transplant rejection. Hyperlipidemia is a significant side effect of sirolimus treatment, and frequently leads to cardiovascular disease. This study was undertaken to determine the repeatability, reversibility, and dose dependence of the plasma lipid and apolipoprotein altering effects of sirolimus, and to elucidate the mechanism by which sirolimus induces hypertriglyceridemia in some renal transplant patients. Six patients with renal allografts maintained on cyclosporine A and prednisone were selected on the basis of their previous hyperlipidemic response to short term (14 days) sirolimus administration. For longer-term treatment, each patient was started on 10 mg/day sirolimus and continued as tolerated for 42 days to reinduce hyperlipidemia. Timed blood samples were analyzed for lipid, apolipoprotein, and sirolimus levels. During sirolimus administration, mean total plasma cholesterol increased from 214 mg/dl to 322 mg/dl (+50%; range 25–92%); LDL-cholesterol levels followed a similar pattern. Mean triglyceride level rose from 227 to 432 mg/dl (+95%; range 9–254%). ApoB-100 concentration rose from 124 to 160 mg/dl (+28%; P < 0.05). ApoC-III level increased from 28.9 to 55.5 mg/dl, +92%; (P < 0.013). These lipid and apolipoprotein changes were found to be repeatable, reversible, and dose dependent. [13C4]palmitate metabolic studies in four patients with hypertriglyceridemia indicated that the free fatty acid pool was expanded by sirolimus treatment (mean = 42.3%). Incorporation of [13C4]palmitate into triglycerides of VLDL, IDL, and LDL was decreased 38.3%, 42,1%, and 38.4%, respectively, by sirolimus treatment of these patients.These results suggest that sirolimus alters the insulin signaling pathway so as to increase adipose tissue lipase activity and/or decrease lipoprotein lipase activity, resulting in increased hepatic synthesis of triglyceride, increased secretion of VLDL, and increased hypertriglyceridemia. Sirolimus (Rapammune®, rapamycin, RAPA) is a potent immunosuppressive drug that reduces renal transplant rejection. Hyperlipidemia is a significant side effect of sirolimus treatment, and frequently leads to cardiovascular disease. This study was undertaken to determine the repeatability, reversibility, and dose dependence of the plasma lipid and apolipoprotein altering effects of sirolimus, and to elucidate the mechanism by which sirolimus induces hypertriglyceridemia in some renal transplant patients. Six patients with renal allografts maintained on cyclosporine A and prednisone were selected on the basis of their previous hyperlipidemic response to short term (14 days) sirolimus administration. For longer-term treatment, each patient was started on 10 mg/day sirolimus and continued as tolerated for 42 days to reinduce hyperlipidemia. Timed blood samples were analyzed for lipid, apolipoprotein, and sirolimus levels. During sirolimus administration, mean total plasma cholesterol increased from 214 mg/dl to 322 mg/dl (+50%; range 25–92%); LDL-cholesterol levels followed a similar pattern. Mean triglyceride level rose from 227 to 432 mg/dl (+95%; range 9–254%). ApoB-100 concentration rose from 124 to 160 mg/dl (+28%; P < 0.05). ApoC-III level increased from 28.9 to 55.5 mg/dl, +92%; (P < 0.013). These lipid and apolipoprotein changes were found to be repeatable, reversible, and dose dependent. [13C4]palmitate metabolic studies in four patients with hypertriglyceridemia indicated that the free fatty acid pool was expanded by sirolimus treatment (mean = 42.3%). Incorporation of [13C4]palmitate into triglycerides of VLDL, IDL, and LDL was decreased 38.3%, 42,1%, and 38.4%, respectively, by sirolimus treatment of these patients. These results suggest that sirolimus alters the insulin signaling pathway so as to increase adipose tissue lipase activity and/or decrease lipoprotein lipase activity, resulting in increased hepatic synthesis of triglyceride, increased secretion of VLDL, and increased hypertriglyceridemia. Sirolimus (Rapammune®, rapamycin, RAPA) is a novel macrocyclic lactone immunosuppressive drug capable of significantly reducing acute graft rejection in kidney (1Kahan B.D. Efficacy of sirolimus compared with azathioprine for reduction of acute allograft rejection: a randomized multicenter study. The Rapamune US Study Group.Lancet. 2000; 356: 194-202Abstract Full Text Full Text PDF PubMed Scopus (862) Google Scholar), liver (2Watson C.J. Friend P.J. Jamieson N.V. Frick T.W. Alexander G. Gimson A.E. Calne R. Sirolimus a potent new immunosuppressive agent for liver transplantation.Transplantation. 1999; 67: 505-509Crossref PubMed Scopus (182) Google Scholar), and heart (3Stroatman L.P. Coles J.G. Pediatric utilization of rapamycin for severe cardiac allograft rejection.Transplantation. 2000; 70: 541-543Crossref PubMed Scopus (27) Google Scholar) transplant patients. Previous studies have shown that sirolimus reduces the incidence of acute rejection when administered in conjunction with cyclosporine and prednisone (1Kahan B.D. Efficacy of sirolimus compared with azathioprine for reduction of acute allograft rejection: a randomized multicenter study. The Rapamune US Study Group.Lancet. 2000; 356: 194-202Abstract Full Text Full Text PDF PubMed Scopus (862) Google Scholar). Furthermore, sirolimus inhibits vascular smooth muscle cell proliferation and reduces neointimal formation in humans, rats, and pigs, thereby attenuating restenosis following angioplasty (4Poon M. Marx S.O. Gallo R. Badimon J.J. Taubman M.B. Marks A.R. Rapamycin inhibits vascular smooth muscle cell migration.J. Clin. Invest. 1996; 98: 2277-2283Crossref PubMed Scopus (463) Google Scholar, 5Burke S.E. Lubbers N.L. Chen Y.W. Hsieh G.C. Mollison K.W. Luly J.R. Wegner C.D. Neointimal formation after balloon-induced vascular injury in Yucatan minipigs is reduced by oral rapamycin.J. Cardiovasc. Pharmacol. 1999; 33: 829-835Crossref PubMed Scopus (107) Google Scholar, 6Gallo R. Padurean A. Jayaraman T. Marx S. Roque M. Adelman S. Chesebro J. Fallon J. Fuster V. Marks A. Badimon J.J. Inhibition of intimal thickening after balloon angioplasty in porcine coronary arteries by targeting regulators of the cell cycle.Circulation. 1999; 99: 2164-2170Crossref PubMed Scopus (481) Google Scholar). Sirolimus binds to the immunophilin FK506 binding protein (FKBP12). Sirolimus/FKBP12 binary complex does not bind to calcineurin, and therefore is not neurotoxic or nephrotoxic. Instead, sirolimus/FKBP12 binds to a protein kinase called mammalian target of rapamycin (mTOR). mTOR controls proteins that regulate mRNA translation initiation and G1 progression (7Brown E.J. Albers M.W. Shin T.B. Ichikawa K. Keith C.T. Lane W. Schreiber S.L. A mammalian protein targeted by G1- arresting rapamycin-receptor complex.Nature. 1994; 369: 756-758Crossref PubMed Scopus (1662) Google Scholar). Recent studies have shown that mTOR directly phosphorylates p70S6 kinase (8Raught B. Gingras A.C. EIF4E activity is regulated at multiple levels.Int. J. Biochem. Cell. Biol. 1999; 31: 43-57Crossref PubMed Scopus (247) Google Scholar), the eukaroytic translation initiator protein 4G1 (eIF4G1), and translation inhibitor (4E-BP1) (8Raught B. Gingras A.C. EIF4E activity is regulated at multiple levels.Int. J. Biochem. Cell. Biol. 1999; 31: 43-57Crossref PubMed Scopus (247) Google Scholar, 9Brunn G.J. Hudson C.C. Sekulic A. Williams J.M. Hosoi H. Houghton P.J. Lawrence J.C. Abraham R.T. Phosphorylation of the translational repressor PHAS-1 by the mammalian target of rapamycin.Science. 1997; 277: 99-101Crossref PubMed Scopus (809) Google Scholar, 10Burnett P.E. Barrow R.K. Cohen N.A. Snyder S.H. Sabatini D.M. RAFT-1 phosphorylation of the translational regulator p70S6 kinase and 4EBP-1.Proc. Natl. Acad. Sci. USA. 1998; 95: 1432-1437Crossref PubMed Scopus (937) Google Scholar). Therefore, inhibition of mTOR by sirolimus contributes to translational arrest by down-regulation of p70S6K, and by increasing the affinity of 4E-BP1 (11Kumar V. Sabatini D. Pandey P. Gingras A.-C. Majumder P.K. Kumar M. Yaun Z.-M. Carmicheal G. Weichselbaum R. Sonenberg N. Kufe D. Kharbanda S. Regulation of the rapamycin and FKBP-target 1/mammalian target of rapamycin and cap-dependent initiation of translation by the c-abl protein-tyrosine kinase.J. Biol. Chem. 2000; 275: 10779-10787Abstract Full Text Full Text PDF PubMed Scopus (50) Google Scholar). Consequently, sirolimus immunosuppressive action is due to inhibition of T-cell activation at a later stage of the cell cycle, G1, and inhibition of p70S6K (10Burnett P.E. Barrow R.K. Cohen N.A. Snyder S.H. Sabatini D.M. RAFT-1 phosphorylation of the translational regulator p70S6 kinase and 4EBP-1.Proc. Natl. Acad. Sci. USA. 1998; 95: 1432-1437Crossref PubMed Scopus (937) Google Scholar). A major adverse reaction associated with sirolimus therapy is hyperlipidemia, a major risk factor for cardiovascular disease, and the most common cause of death after renal transplantation (12United States Renal Data System (USRDA). 1998. The National Institute of Health, National Institute of Diabetes, and Digestive and Kidney Disease. Annual Data report, Bethesda, Maryland.Google Scholar). Several studies have shown an increase in serum triglyceride levels in renal transplant recipients treated with sirolimus (13Brattström C. Wilczek H.E. Tydén G. Böttiger Y. Säwe J. Groth C.G. Hypertriglyceridemia in renal transplant recipients treated with sirolimus.Transplant. Proc. 1998; 30: 3950-3951Crossref PubMed Scopus (45) Google Scholar, 14Brattström C. Wilczek H. Tyden G. Böttiger Y. Säwe J. Groth C.G. Hyperlipidemia in renal transplant recipients treated with Sirolimus (Rapamycin).Transplantation. 1998; 65: 1273-1274Google Scholar). Their hyperlipidemia was dose-dependent and reversible within 1 to 2 months after discontinuation of treatment (13Brattström C. Wilczek H.E. Tydén G. Böttiger Y. Säwe J. Groth C.G. Hypertriglyceridemia in renal transplant recipients treated with sirolimus.Transplant. Proc. 1998; 30: 3950-3951Crossref PubMed Scopus (45) Google Scholar, 14Brattström C. Wilczek H. Tyden G. Böttiger Y. Säwe J. Groth C.G. Hyperlipidemia in renal transplant recipients treated with Sirolimus (Rapamycin).Transplantation. 1998; 65: 1273-1274Google Scholar). In the present study, we have examined the dependence of lipid, lipoprotein, and apolipoprotein levels, as well as fatty acid and triglyceride metabolism on sirolimus dosage and treatment duration in renal allograft recipients with different types of hyperlipidemia. This study was performed in six patients, each of whom had received a renal allograft within 3–8 years at Hermann Hospital Transplant Center, Houston, Texas. These patients were selected based on their previous hyperlipidemic response upon short-term treatment (14 days) with sirolimus and had stable renal allografts. All patients selected had no evidence of hepatic or biliary dysfunction, as reflected in serum transaminase levels not more than 20% above normal limits, nor lipid abnormalities with triglycerides >400 mg/dl or cholesterol >250 mg/dl. Patients indicated their willingness to participate in the study by signing a consent form, approved by the Institutional Review Boards for human research at the University of Texas Health Science Center-Houston and Baylor College of Medicine and its affiliated hospitals. The patient group included four females and two males aged 27–55 years (44.8 ± 10.6) (Table 1). All six patients were maintained on cyclosporine A (CsA) (Neoral), prednisone, and diuretic therapy. Four patients had developed mixed hyperlipidemia, one had developed hypercholesterolemia, and one had developed hypertriglyceridemia in response to the previous short-term sirolimus treatment (Table 2). Patients who had been diagnosed with diabetes were receiving lipid-lowering medications (Table 1). This regimen was continued unaltered while on sirolimus during the longer 6-week study (42 days). All six patients had the apo-E3/E3 genotype.TABLE 1.Clinical and demographic data of the renal transplant re-challenge (42 days) study participantsIDAgeGenderRaceDonor TypeLipid RXNeoral (N) DosePrednisone DoseCreatinine Before SirolimusCreatinine After SirolimusDiabetesDiagnosismgmg/dlWAS-1/227FHispanicLRDNone100/75aaDose changed to 75/50 by the end of sirolimus treatment.7.51.72.5NoESRD/GNWAS-3/455FBlackCadP125/1007.50.90.9IDDM/SIESRD/HTNWAS-5/653FHispanicLRDL, P125/12550.70.6NIDDMESRD/HTNWAS-7/849FHispanicLRDL, P125/1007.51.51.1IDDM/SIESRD/CPWAS-9/1037MWhiteLRDNone100/10051.73.1NoESRD/LupusWAS-11/1248MHispanicLRDL, P200/1757.51.61.7NIDDMESRD/HTNLRD, live related donor; P, pravacol; L, lopid; ESRD, end stage renal disease; GN, gynecological disease; HTN, hypertension; CP, chronic pyelonephritis; IDDM/SI, insulin dependent diabetes mellitus/steroid induced; Cad, cadaveric.a aDose changed to 75/50 by the end of sirolimus treatment. Open table in a new tab TABLE 2.Plasma cholesterol and trigylceride response of renal transplant recipients to the initial shorter term sirolimus treatment (14 days)Cholesterol Level Before/ After SirolimusTriglyceride Level Before/ After SirolimusHyperlipidemiaPatient IDSirolimus DoseDay −1bbSirolimus treatment was started one day after renal transplantation.Day 14 Stop DrugDay 28Day 56ChangeDay −1bbSirolimus treatment was started one day after renal transplantation.Day 14 Stop DrugDay 28Day 56ChangeBeforeAftermg/daymg/dl%mg/dl%WAS-1/22264416aaAt this time, patient WAS-1/2 was on sirolimus treatment for 61 days.NDND57169465aaAt this time, patient WAS-1/2 was on sirolimus treatment for 61 days.NDND175II aII bWAS-3/41294246337300−16123294211163139II aII bWAS-5/66249373293ND497391709652ND131II bII bWAS-7/8633339929325220248628313155153II bII bWAS-9/10714824119118062156340162137118NII bWAS-11/122318310341303−331539247529824II bII bMean28123P value0.10.04ND, not determined. Percent change is the difference between day –1 and day 14 (when sirolimus treatment was stopped).a aAt this time, patient WAS-1/2 was on sirolimus treatment for 61 days.b bSirolimus treatment was started one day after renal transplantation. Open table in a new tab LRD, live related donor; P, pravacol; L, lopid; ESRD, end stage renal disease; GN, gynecological disease; HTN, hypertension; CP, chronic pyelonephritis; IDDM/SI, insulin dependent diabetes mellitus/steroid induced; Cad, cadaveric. ND, not determined. Percent change is the difference between day –1 and day 14 (when sirolimus treatment was stopped). Two weeks before commencing the study, each candidate patient underwent screening evaluation at the Hermann Hospital Transplant Center. A complete physical examination and a series of biochemical screening tests were performed to determine suitability for the study. Blood sampling for the lipid profile and lipolytic enzymes (e.g., post heparin lipase) were scheduled to avoid interference of one test with another. The initial shorter-term study (14 days) was conducted immediately after transplantation. Each patient was treated with a constant dose (1–7 mg/day) of sirolimus over a 14 day period (Table 2). For the longer-term study (42 days), qualifying patients began the 8-week protocol on day −6 with a pre-drug lipoprotein metabolic study lasting 6 days, up to day 1. On day 1, each patient started the sirolimus treatment initially at a level of 10 mg/day for 42 days. While on sirolimus, the patient returned weekly or biweekly to the outpatient center for determination of lipids, lipoproteins, apolipoproteins, lipid enzymes, and sirolimus trough levels. If the patient's lipid levels exceeded an acceptable range, then the sirolimus dose was reduced as described previously (15Hoogeveen R.C. Ballantyne C.M. Pownall H.J. Opekun A.R. Hachey D.L. Jaffe J.S. Oppermann S. Kahan B.D. Morrisett J.D. Effect of rapamycin on the metabolism of Apo B-100 containing lipoproteins in renal transplant patients.Transplantation. 2001; 72: 1244-1250Crossref PubMed Scopus (117) Google Scholar). After 42 days on treatment, a second lipoprotein metabolic study was initiated, lasting 6 days until day 47, after which sirolimus treatment was discontinued. Cyclosporin and prednisone maintenance therapy were continued. Each patient returned to the outpatient center on day 56 to give another fasting follow-up blood sample for determination of the final lipid and lipoprotein profile, and to undergo the closeout physical examination. Sirolimus trough levels were measured on whole blood samples with a multi-step liquid-liquid extraction followed by reversed-phase-HPLC with ultraviolet detection performed by Dr. Kim Napoli at the Organ Transplantation Center of the University of Texas Health Science Center at Houston (16Napoli K.L. Kahan B.D. Routine clinical monitoring of sirolimus (rapamycin) whole-blood concentrations by HPLC with ultraviolet detection.Clin. Chem. 1996; 42: 1944Crossref Scopus (82) Google Scholar). Lipid, lipoprotein, and apolipoprotein measurements were performed in the Atherosclerosis Lipid Laboratory of The Methodist Hospital. Plasma samples were prepared by centrifugation (1500 g, 10 min, 4°C) of venous blood collected after 12 h fasting into Vacuutainertm tubes containing EDTA. Total plasma cholesterol (17Seidel J. Hagele E. Ziegenhorn J. Wahlenfeld J. Reagents for the enzymatic determination of serum total cholesterol with improved lipolytic efficiency.Clin. Chem. 1983; 29: 1075-1080Crossref PubMed Scopus (877) Google Scholar) and triglycerides (18Nagele U. Hagele E. Sauer G. Wiedermann E. Lehmann P. Wahlefeld A.W. Gruber W. Reagent for the enzymatic determination of serum total triglyceride with improved lipolytic efficacy.J. Clin. Chem. Clin. Biochem. 1984; 22: 165-174PubMed Google Scholar) were measured enzymatically (Boehringer Mannheim Diagnostics). LDL cholesterol (LDL-C) levels were determined directly from the plasma after immunoprecipitation of VLDL and HDL using a kit from Sigma Chemical Co. (St. Louis, MO). HDL-C was determined by measuring cholesterol in the supernatant liquid after precipitation of the VLDL and LDL with MgCl2 and dextran sulfate (19Warnick G.R. Benderson J. Albers J.J. Dextran Sulfate-Mg2+ precipitation procedure for the quantification of high density-lipoprotein cholesterol.Clin. Chem. 1982; 28: 1379-1388Crossref PubMed Scopus (1812) Google Scholar). Plasma apoB-100 was measured by ELISA using Mab RP-066 (Intracel, Inc., Rockville, MD). ApoA-I was measured by nephelometry of the precipitate formed with anti-apoA-I (IncStar, Inc.). ApoC-II, and apoC-III were determined by radial immunodiffusion (Daichi, Ltd.). ApoE genotyping was performed using a PCR based method (20Hixson J.E. Vernier D.T. Restriction isotyping of the human apolipoprotein E by gene amplification and cleavage with Hha1.J. Lipid Res. 1990; 31: 545-548Abstract Full Text PDF PubMed Google Scholar). Patients were fasted overnight prior to the start of their metabolic studies (15Hoogeveen R.C. Ballantyne C.M. Pownall H.J. Opekun A.R. Hachey D.L. Jaffe J.S. Oppermann S. Kahan B.D. Morrisett J.D. Effect of rapamycin on the metabolism of Apo B-100 containing lipoproteins in renal transplant patients.Transplantation. 2001; 72: 1244-1250Crossref PubMed Scopus (117) Google Scholar). Sodium [13C 4]palmitate (Isotec, Inc., Miamisburg, OH) complexed to human serum albumin (Centeon, LLC. Kankakee, IL) was administered by constant intravenous infusion (0.6 mg/kg/h) over 7 h. Each patient was given oral Sustecal (30 kcal/kg), which was consumed in 16 equal portions at hourly intervals, providing 22% of calories from fat and 0.88 g protein/kg. Blood samples (15 ml) were drawn, 18 over the first 24 h and 1 daily for the next 5 days, from which VLDL, IDL, and LDL were isolated by density gradient ultracentrifugation (21Redgrave T.G. Roberts D.C.K. West C.E. Separation of plasma lipoproteins by density gradient ultracentrifugation.Anal. Biochem. 1975; 65: 42-49Crossref PubMed Scopus (870) Google Scholar). These lipoproteins were delipidated by organic solvent extraction (CHCl3-CH3OH, 2:1, v/v) and the different lipid fractions separated by thin layer chromatography (22Skipski V.P. Barclay M. Thin Layer Chromatography of Lipids (Ed: J.M. Lowenstein).Methods Enzymol. 1969; XIV: 530-598Crossref Scopus (476) Google Scholar). The triglyceride fraction was hydrolyzed with 15% KOH and derivatized with pentafluorobenzylbromide (23Hachey D.L. Patterson B.W. Elsas L.J. Reeds P.J. Klein P.D. Isotopic determination of organic keto acid pentafluorobenzyl esters in biological fluids by negative chemical ionization gas chromatography/ mass spectrometry.Anal. Chem. 1991; 63: 919-923Crossref PubMed Scopus (79) Google Scholar). Plasma free fatty acids (FFAs) were isolated by solid phase extraction columns (Alltech Associates, Inc, Deerfield, IL) and derivatized by the same methodology as used for triglycerides. The resulting pentafluorobenzyl (PFB) ester (m/z 259) was analyzed for [13C4]palmitate enrichment by gas chromatography (Supelco fused silica capillary column, 30 m × 0.25 mm × 0.25 μm film) and mass spectrometry (Hewlett Packard HP 6890GC/ 5973 MSD). The effects of sirolimus treatment on fasting lipids and lipoprotein levels were evaluated using Student's paired t-tests (24Matthews D.E. Farewell V.T. Using and understanding Medical Statistics. 2nd edition. S. Karger AG, Basel, Switzerland1988: 113-119Google Scholar). Log or rank transformations were utilized when needed to meet the assumptions of the t-test. Statistical analyses were conducted using STATA (Release 4.0) and Prism (version 2.0) software. Plots of percent atom enrichment before and after sirolimus treatment were quantitatively compared by calculating the area under the curve out to 36 h. The first and last data points were used to determine the baseline amplitude, which was used to correct the integrated area. A major objective of the current study was to determine if sirolimus-induced hyperlipidemia in renal transplant patients is reproducible, reversible, and dose-dependent. Although the shorter-term study (14 days), conducted immediately after transplantation, suggested reversibility of the effect (Table 2), the measurements did not include a complete lipoprotein and apolipoprotein profile, nor were the measurements frequent enough to closely monitor sirolimus-induced changes. Furthermore, in that initial study, each patient was treated with a constant dose (1–7 mg/day) over a 14 day period (Table 2), whereas in the later longer term study (42 days) all patients were started at 10 mg/day with the express purpose of re-inducing hyperlipidemia (Table 3). As anticipated, it was necessary to reduce the dose in those patients who exhibited sirolimus-induced hyperlipidemia exceeding the level allowed by the protocol. Dosage was also reduced if required by blood chemistries or cell count, and to bring the elevated creatinine values to normal levels. These dosage adjustments usually prevented the patients from achieving constant trough levels of sirolimus, but they enabled the observation of dose-dependent lipid changes that would not have been detected with a constant dose strategy. Monitoring sirolimus blood concentrations revealed that drug trough levels reflected dose in all patients except Wyeth Ayerst study patient before/after sirolimus treatment (WAS-11/12), whose concentrations steadily decreased despite continuous high dosing (10 mg/day) throughout the study (Fig. 1A–F, all parts labeled II).TABLE 3.Plasma cholesterol and trigylceride response of renal transplant recipients to re-challenge with sirolimus treatment (42 days)Cholesterol Level Before/ After SirolimusTriglyceride Level Before/ After SirolimusHyperlipidemiaPatient IDDay −1Day 42 Stop DrugChangeDay −1Day 42 Stop DrugChangeBeforeAftermg/dl%mg/dl%WAS-1/2244431+92275974+254II bII bWAS-3/4273366+34225284+26II aII bWAS-5/6234301+29376491+31II bII bWAS-7/8224394+75165481+191II aII bWAS-9/10131216+65135212+57NII aWAS-11/12177222+25140aaSince the Day 1 point appeared spurious, the value used for this patient was the average of Day –7 and Day +7 value.153+9NII aMean213.4321.7+50227432+95S.D.519087299P value0.0070.1a aSince the Day 1 point appeared spurious, the value used for this patient was the average of Day –7 and Day +7 value. Open table in a new tab In the initial, shorter-term study, sirolimus caused variable changes in total cholesterol levels within 14–28 days (range: −3 to 62%; mean: +28%; Table 2). In the later 6-week study (42 days), sirolimus caused marked increases in the total cholesterol levels in five of the six patients (range: +25 to +92%; mean: +50%; P = 0.007; Table 3). The longer duration and larger dosage are likely reasons for the greater elevation of total cholesterol in the second study. Frequent measurements during the second study indicated rather gradual increases in cholesterol levels (Fig. 1A–F, all parts labeled II) from the time the drug was started (day 1) to the time it was stopped (day 42). Because triglyceride levels often exceeded the 400 mg/dl limit for which the Friedewald equation is valid for calculating LDL-C, it was necessary to measure this analyte directly (dLDL-C). In general, dLDL-C increased gradually with sirolimus treatment, resembling the changes seen in total cholesterol with respect to timing but not magnitude (Fig. 1A–F, all parts labeled II). Throughout the entire 6-week treatment period (42 days), sirolimus had no effect on HDL-C levels any of the patients besides WAS-3/4. This patient had a remarkably high initial HDL-C level (92 mg/dl), which rose slowly and monotonically, reaching a plateau level of 110 mg/dl at day 28 (Fig. 1A, part II). In the initial shorter-term study (Table 2), sirolimus induced a substantial increase in the triglyceride levels of every patient (range: +24 to 175%; mean: +123%). For the later longer-term study (Table 3), sirolimus again elevated triglyceride levels (range: +9 to 254%; mean: +95%), but these changes were not as great as those observed in the short term study. Only two patients, WAS-1/2 and WAS-7/8, had elevations greater in the longer-term than the shorter-term study (Tables 2 and 3). In general, triglyceride levels were highly responsive to sirolimus dosage. This is well illustrated in responses of patients WAS-3/4 and WAS-9/10, in which the initial 10 mg/day induced a rapid rise in triglyceride levels; necessary reductions in dosage resulted in prompt reductions in triglyceride, and subsequent increments of dosage induced yet a second set of increases in triglycerides. The single exception to these observations was seen in patient WAS-11/12, who received a full 10 mg/day dose but maintained a comparatively stable triglyceride level (range: 110–210 mg/dl) throughout the 42 day treatment period. The apolipoprotein showing the greatest response to sirolimus was apoB-100, a major protein component of VLDL and LDL. Hence, its dose dependent changes reflect the composite changes in triglyceride (transported primarily by VLDL) and cholesterol (transported primarily by LDL). This point is illustrated in apoB-100 levels of patient WAS-9/10 that rose to a maximum of 120 mg/dl at day 14, corresponding to the maximum triglyceride level of this patient (410 mg/dl) at the same day. The abrupt decrease in triglyceride to 105 mg/dl at Day 28 is somewhat attenuated in the apoB-100 curve, due in part to the much slower decrement in LDL-C (Fig. 1E, parts II and III). ApoC-II and apoC-III are important protein components of VLDL and HDL. The plasma levels of apoC-II were typically low and did not change appreciably during the course of the study (range: −2.4 to +10.6 mg/dl; mean: 3.25 mg/dl, P = 0.18). In contrast, the initial values of apoC-III were substantial, and increased significantly between day 1 and day 42 (range: +7 to + 53 mg/dl; mean 27 mg/dl; P = 0.013). Since apoC-II is an activator (25Bengtsson G. Olivecrona T. Lipoprotein lipase: some effects of activator proteins.Eur. J. Biochem. 1980; 106: 549-555Crossref PubMed Scopus (96) Google Scholar) and apoC-III is an inhibitor (26McConathy W.J. Gesquiere J.C. Bass H. Tartan A. Fruchart J.C. Wang C.S. Inhibition of lipoprotein lipase activity by synthetic peptides of apolipoprotein CIII.J. Lipid Res. 1992; 33: 995-1003Abstract Full Text PDF PubMed Google Scholar, 27Ginsberg H.N. Le N.A.A. Goldberg I.J. Gibson J.C. Rubinstein A. Wang-Iverson P. Norum R. Brown W.V. Apolipoprotein B metabolism in subjects with deficiency of apo CIII and AI. Evidence that apolipoprotein CIII inhibits catabolism of triglyceride rich lipoproteins by lipoprotein lipase in vivo.J. Clin. Invest. 1986; 78: 1287-1295Crossref PubMed Scopus (339) Google Scholar) of lipoprotein lipase (LPL), these results provide a reasonable explanation for the substantially lower LPL activity in these renal transplant patients (∼20–70%) compared with normolipidemic controls (15Hoogeveen R.C. Ballantyne C.M. Pownall H.J. Opekun A.R. Hachey D.L. Jaffe J.S. Oppermann S. Kahan B.D. Morrisett J.D. Effect of rapamycin on the metabolism of Apo B-100 containing lipoproteins in renal transplant patients.Transplantation. 2001; 72: 1244-1250Crossref PubMed Scopus (117) Google Scholar). ApoA-I is a principal apolipoprotein component of HDL. In patients WAS-1/2, -5/6, -7/8, and -11/12, apoA-I levels were not remarkably affected by sirolimus treatment. However, in patient WAS-3/4, apoA-I rose 58% (Fig. 1B, part III), and in patient WAS-9/10 it rose 50% (Fig. 1E, part III) over the 42 day treatment period. The increase in apoA-I was attended by an increase in HDL for patient WAS-3/4 (Fig. 1B, part II) but not for patient WAS-9/10 (Fig. 1E, part II). After 42
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