RAD(40-O-[2-hydroxyethyl]-rapamycin) (Novartis Pharmaceuticals Corp., EastHanover, NJ) is a novel macrolide with potent immunosuppressive andantiproliferative properties that is currently being evaluated in clinicaltrials for prevention of solid organ allotransplant rejection. RAD, astructural analogue of sirolimus, displays distinctive pharmacokineticcharacteristics: a shorter half-life and less time to attain steady stateconcentrations. Distinct from the calcineurin inhibitors, cyclosporine andtacrolimus, RAD acts at a later stage of the T-cell response to alloantigensby blocking transduction of signals generated by growth factors, such asinterleukin-2 (1).The complementary actions of RAD and cyclosporine provide the basis for asynergistic interaction, which has been demonstrated in vitro by usinglymphocyte proliferation assays and in vivo in rat transplantation models (2).The inhibitory effect of RAD is not restricted to T cells; it also inhibitsthe signals provided by an array of hematopoietic and nonhematopoieticcell-growth factors, including those that stimulate vascular smooth musclecells (1).Based on RAD’s mode of action (1),acceptable tolerability profile (3, 4),and potential synergistic effect with cyclosporine (2),this study was initiated to evaluate the effects of RAD in de novo renaltransplant recipients. This dose-ranging study assessed the incidence andseverity of acute renal rejection episodes as well as the tolerability of 3doses of RAD in combination with Neoral® andcorticosteroids.
Maleand female de novo renal transplant recipients, 16 to 65 years of age, wereeligible for enrollment at 8 transplant centers in the United States, Canada,and Europe. Recipients of primary cadaveric, living-unrelated, ornon-HLA-identical, living-related donor kidney transplants were eligible forinclusion if cold ischemic time was <40 hr and if they had not receivedantibody induction or investigational drug therapy within the 4 weekspreceding randomization. Recipients who displayed uncontrolledhypercholesterolemia (≥350 mg/dL), hypertriglyceridemia (≥750 mg/dL), awhite blood cell count ≤4×10 9 /L, anabsolute neutrophil count ≤2×10 9 /L, aplatelet count ≤100×10 9 /L, severe systemicinfections, past or present malignancy, or coagulopathy were excluded from thestudy. Female patients of childbearing potential were required to have anegative serum pregnancy test before study enrollment and to practice anapproved method of birth control during the study. All enrollees were requiredto sign an informed consent document approved by the local humansubject’s committee. The study was conducted according to rules of GoodClinical Practice as defined by the United States Code of Federal Regulations#21, the European Directive 91/507/EEC, and the Declaration ofHelsinki.
This ongoing, 1-year, multicenter, randomized,double-blind, parallel group, dose-ranging study in de novo renal transplantrecipients compared the effects of 1 mg, 2 mg, or 4 mg of RAD administereddaily in two divided doses in combination with Neoral® (cyclosporine, USPMODIFIED, Novartis) and corticosteroids. The results reported here representthe primary analysis stipulated to be performed at 6 months. After theinvestigator determined that the graft was functional, recipients wererandomized to one of three RAD dosage groups. Study medication was begun onthe day of randomization and within 48 hr posttransplant. Neoral® wasadministered initially at 6–12 mg/kg/day with subsequent doses adjustedto maintain the following 12 hr target trough levels: weeks 1–4,150–400 ng/ml; months 2–6, 75–300 ng/ml. After perioperativeadministration of methylprednisolone, corticosteroids were taperedpostoperatively according to each center’s protocol but maintained aminimum dosage equivalent to 5 mg/day of prednisone for at least 6 months. Noimmunosuppressive drugs other than those specified by the protocol wereallowed. All patients were mandated to receive prophylaxis for Pneumocystis carinii pneumonia. Insituations of donor-positive and recipient-negative transplants,cytomegalovirus prophylaxis was administered according to center practice.Lipid-lowering therapy was prescribed by the investigator, as needed. Dosereduction or interruption of study medication was allowed in cases ofdiminished platelet (<75×10 9 /L) or whiteblood cell (<3×10 9 /L) counts or at theinvestigator’s discretion as dictated by a moderate or severe adverseevent.
The protocol stipulated that recipientsexperiencing a suspected rejection episode undergo a transplant biopsy, whichwas scored by the local pathologist using the Banff criteria (5).Mild to moderate rejection episodes were to be treated initially with500–1000 mg of intravenous methylprednisolone for 1–5 days.Administration of antilymphocyte antibody according to center-specificpractice was permitted for corticosteroid-resistant or vascular rejectionepisodes.
Eligibletransplant recipients were evaluated at baseline (within the first 48 hrposttransplantation and before randomization), once a week for the first 4weeks and at months 2, 3, and 6. Follow-up information was requested at 3 and6 months in the event that study medication was prematurely discontinued.Efficacy was assessed according to the incidence and severity of biopsy-provenacute rejection episodes, graft survival, and patient survival. Safety wasassessed by electrocardiography, vital signs (radial pulse rate, bloodpressure, weight), laboratory tests (hematology, urinalysis, biochemistry),and the occurrence of treatment-emergent adverseevents.
The population size of approximately 100 patients(33 in each dosage group) was selected to provide a reasonable number ofpatients per group to assess tolerability and was not based on determinationsof the power to detect statistically significant intergroup differences inefficacy or safety outcomes. Therefore, results of statistical analyses areconsidered descriptive only.
Demographic information wassummarized by frequency distributions (for categorical variables) anddescriptive statistics, including mean and standard deviation (for continuousvariables). Continuous variables were compared between the treatment groups byusing a Kruskal-Wallis test and categorical variables with Pearson’schi-square test.
All efficacy and safety analyses wereperformed by using the intent-to-treat principle, i.e., all randomizedpatients who received at least one dose of RAD were included. Only datacollected within 6 months (visit window out to 225 days) from the first dosewere included in this analysis. Kaplan-Meier estimates for the rate ofbiopsy-proven acute rejection episodes were determined for all three treatmentgroups. Any patient who prematurely discontinued the study was censored in theKaplan-Meier estimates of rejection rates. Pearson’s chi-square test wasused to compare the percentages of patients with biopsy-proven acute rejectionepisodes, patients with rejections requiring antibody treatment, patientsurvival and graft survival, and a composite efficacy failure variable thatincluded death, graft loss, biopsy-proven acute rejection episodes, andpatients lost to follow-up between treatment groups. The percentage ofpatients with moderate-to-severe acute rejection episodes was compared betweengroups using the Fisher’s exact test.
Adverseevents, clinical laboratory test values, and vital sign determinations weresummarized by using descriptive statistics by treatment group for all patientswho were randomized and had reported at least one adverse event or had onesafety measurement. The number of patients with RAD dose alterations due toadverse events was summarized by treatment group, as were the frequencies ofviral and fungal opportunistic infections. For patients who discontinuedtreatment, adverse events reported within 8 days of discontinuation wereincluded. Descriptive statistics, such as mean and standard deviation, werecalculated for vital signs and laboratory variables. Comparisons between thethree treatment groups were made with the Kruskal-Wallis test at eachstipulated visitmilestone.
Demographicsand Patient Disposition
A total of 103 transplant recipientsin the United States (n=57), Germany (n=30), Canada(n=10), and the United Kingdom (n=6) were randomized among the 3RAD treatment groups. Demographic characteristics were similar between groups,except that the percentage of patients with diabetes mellitus was at leasttwice as great in the 1-mg/day group as in the other groups(Table 1).
Sixty-fiverecipients remained on study medication for 6 months. Six recipients in the1-mg/day group, 5 in the 2-mg/day group, and 11 in the 4-mg/day groupdiscontinued treatment due to adverse events or abnormal laboratory values.Four recipients each in the 1-mg/day treatment group (11.8%) and the 4-mg/daygroup (11.4%) discontinued the study medication on account of unsatisfactorytherapeutic effect. Three recipients died, three withdrew consent, and twoother patients were withdrawn from study medication because of protocolviolations. Follow-up information was available for all except 1 recipient (inthe 1-mg/day group). This case was categorized as a treatment failure for thecompositeendpoint.
Efficacyoutcome measures for each treatment group are presented in Table 2. Althoughthe study was not designed to detect statistically significant differences inefficacy outcomes, patients in the 2-mg or 4-mg treatment groups experiencedreductions in the occurrence of biopsy-proven acute rejection episodescompared with those in the 1-mg/day group, namely, 14.7% or 25.7% vs. 32.4%,respectively. Figure 1 shows the Kaplan-Meier estimates of the probability offreedom from biopsy-proven acute rejection episodes for each dose group fromthe time of initiation of therapy until the 6-month time-point. Although thedifferences were not significant, the 2-mg/day and 4-mg/day groups experiencedgreater freedom from rejection episodes than the 1-mg/day group. The majorityof biopsy-proven acute rejection episodes occurred during the first 3 monthsof treatment. Patients in the 1-mg/day group experienced more repeat acuterejection episodes, but no patient experienced more than a total of 2episodes.
Acuterejection episodes among patients receiving either the 2-mg or 4-mg/day dosewere mild, except for one moderate acute rejection episode reported in the4-mg/day group (Table 3). In both the 2-mg/day(P =0.002) and 4-mg/day(P =0.006) groups, significantlyfewer recipients had moderate-to-severe rejection episodes than those in the1-mg/day group (Fig. 2).
Onepatient in each cohort experienced graft loss; in the 1-mg/day group, the losswas due to chronic rejection, in the primary 2-mg/day group, to nonfunction,and in the 4-mg/day group, to progression of rejection after reduction ofimmunosuppression due to intercurrent meningitis, producing death-censoredsurvival rates of 97.1% in all dose arms. Three recipients died withfunctioning renal grafts: 1 (receiving 1 mg/day) due to abdominalischemia/septic shock on day 4; 1 (receiving 2 mg/day) due to perforatedduodenal ulcer and related complications on day 137; and 1 (also receiving 2mg/day) due to myocardial infarction (MI) on day 139. The latter recipient hada history of ischemic heart disease and previously documented MI. No deathswere related to the study medication as judged by the investigators. Theoverall graft survival rates were 94.1%, 91.2%, and 97.1% for the 1-, 2-, and4-mg/day dose arms, respectively.
The number of adverse events, considered tobe drug-related by the investigators, was lowest in the 1-mg/day group. Theincidence of serious adverse events (fatal or life-threatening, requiring orprolonging hospitalization, significantly disabling, constituting cancer orbirth defects, or resulting from overdose) was highest in the 4-mg/day group,and a greater number of patients in this group required reduction,interruption, or discontinuation of RAD therapy due to adverse events(Table 4). Studymedication was discontinued at similar rates in the 3 groups(Table 5). Causesfor discontinuation included pancytopenia (n=1, 1-mg/day group),leukopenia (n=2, 1-mg/day group and 4-mg/day group), thrombocytopenia(n=1, 4-mg/day group), thrombocytopenia and leukopenia (n=1,4-mg/day group), infections (n=7; 2, 3, and 2 in the 1-, 2-, and4-mg/day groups, respectively), death (n=3, 1 in the 1-mg/day group and2 in the 2-mg/day group), pure graft loss (n=1), pancreatitis(n=1), hemolytic uremic syndrome (n=1), and other reasons(n=11). Additionally, 1 patient in the 1-mg/day group discontinuedstudy medication due to an elevated serum-alanine-aminotransferase level,which was considered not related to the study drug. There was 1 malignancy, alymphoma discovered at autopsy in the patient who died of perforated duodenalulcer and related complications on day 137.
Although the overall rate of herpes simplexinfections was low, there was an increased incidence in the 4-mg/day group(Table 6).Cytomegalovirus infections were documented in less than 3% of patients, andmoniliasis constituted the majority of the fungal infections. There were no P. carinii infections.
Changesin Laboratory Values
Platelet and white blood cell counts,serum triglyceride, and cholesterol levels are presented in Table 7. Althoughthe occurrence of thrombocytopenia(<75×10 9 /L) was infrequent (<5% ofoverall patients), the incidence was higher in the 4-mg/day dose group.Similarly, the incidence of leukopenia(<3.0×10 9 /L) was low but more common inthe 4-mg/day group.
The timecourse of triglyceride and cholesterol levels during the 6-month period ispresented in Figure 3, A andB. Mean and mean-maximum triglyceride, as well asmean-maximum cholesterol levels, at 6 months were lowest in the 1-mg/day group(Table 7). Patientswho were treated with hydroxymethyl glutaryl (HMG)-CoA reductase inhibitortherapy displayed higher mean levels of cholesterol at 1 month but showedpositive responses to therapy, resulting in mean cholesterol values at 6months comparable to those of untreated patients(Fig. 3C).Similarly, patients with elevated triglyceride levels showed a positivetherapeutic response to lipid-lowering therapy. The percentage of patientsreceiving lipid-lowering drugs in the 1-, 2-, and 4-mg/day groups was 35%,71%, and 63%, respectively, with 32%, 65%, and 54% of patients, respectively,receiving HMG-CoA reductase inhibitors.
Themean serum creatinine levels during the study were similar among the threegroups. Mean values at 6 months were 1.8 mg/dL, 2.1 mg/dL, and 2.0 mg/dL forthe 1-, 2-, and 4-mg/day dose groups, respectively. No significant differenceswere found between the groups with respect to mean change from nadir toendpoint. No significant differences in changes in mean blood pressure frombaseline to 6 months were observed for either systolic or diastolic bloodpressures.
Mean RAD trough concentrations at 6 monthsfor the 1-, 2-, and 4-mg/day groups were 1.9, 5.3, and 8.4 ng/ml,respectively, reflecting the dose-linear kinetics seen in earlier trials. Meancyclosporine trough levels were not statistically different among groups atany time and at 6 months were 153, 154, and 135 ng/ml,respectively.
Inthis first assessment in de novo renal transplant recipients, RAD administeredin combination with Neoral® and corticosteroids resulted in excellentpatient and graft survival. None of the three deaths was judged by theinvestigators to be related to RAD administration. Among the 103 patients,only 3 graft losses occurred within the first 6 months posttransplant.Furthermore, the immunosuppressive effects of RAD were evidenced by the trendtoward a reduction in the rate of biopsy-proven acute rejection episodes amongpatients treated with 2 or 4 mg/day as compared with the 1-mg/day group. The14.7% incidence of biopsy-proven acute rejection episodes observed amongpatients in the 2-mg/day dose group compares favorably to those reported fromrecent trials of new drugs in combination with cyclosporine andcorticosteroids: 17–25% for sirolimus at 2 mg/day (6) and 17–20% for mycophenolate mofetil (7–9).The unexpected, albeit not statistically significant, difference in acuterejection rates between the 2-mg and 4-mg RAD groups presumably reflects thelimited sample size.
Even more compelling is the findingthat, despite both the absence of a placebo-control group and the relativelysmall size of this dose-ranging study, the severity of acute rejectionepisodes was significantly reduced in the 2-mg/day(P =0.002) and 4-mg/day(P =0.006) groups as comparedwith the 1-mg/day group. This finding may have considerable clinicalimportance, because high grades of acute rejection have been associated withgreater difficulty in achieving full reversal and an increased risk ofdevelopment of chronic allograft nephropathy (10, 11).RAD, by reducing the severity of acute rejection episodes and by inhibitinggrowth factor-induced proliferation, has the potential to reduce the incidenceof chronic allograft nephropathy (1, 12, 13).
Earlierstudies have indicated that high doses of RAD may be associated with decreasesin white blood cell and platelet counts and increases in lipids (3, 4).The results from this study show that these potential side effects do notpreclude the use of RAD in de novo renal transplantation. Significantleukopenia (<3.0×10 9 /L) was observeduncommonly and clinically meaningful platelet level reductions(<75×10 9 /L) were also uncommon (5patients), occurring primarily at the 4-mg/day dose (4 patients). Thehyperlipidemia observed in all cohorts is clearly multifactorial, becauseelevations of serum cholesterol and triglyceride levels have been reported inposttransplant patients receiving only cyclosporine and corticosteroids withor without azathioprine (14).The observations that no patients discontinued the study drug for this reason,and that these elevations responded to lipid-lowering therapy, suggest thatthese changes are clinically manageable by steroid dose reduction andcountermeasure therapy, including dietary intervention and lipid-loweringagents. In fact, mean cholesterol levels at 6 months were similar among allRAD groups and comparable to those observed in patients treated withcyclosporine and corticosteroids (with or without azathioprine) in the controlarms of two sirolimus trials (224 and 236 mg/dL) (6).At 6 months, mean serum triglyceride levels in the 1-mg/day RAD group werecomparable to those treated with cyclosporine and steroids alone. Both the 2-and 4-mg/day RAD groups had similar or lower triglyceride levels than the2-mg/day sirolimus dosage group (6).
Despitethe reduction in the incidence and severity of acute rejection episodes in the2-mg/day group, the incidence of opportunistic infections (viral plus fungal)was similar to that seen in the 1-mg/day group. Patients in the 4-mg/day groupexperienced more viral or fungal infections and more discontinuations fromstudy drug. Overall, these findings suggest that the optimal dose is between 1mg and 4 mg/day.
In addition to the positive clinicalfindings with RAD in this initial trial in de novo renal transplantation, arecent report has pointed to additional potential benefits of RAD. Unlikecyclosporine, tacrolimus, and sirolimus, which inhibit high-energy phosphatemetabolism in rat-brain slices in vitro, RAD did not inhibit this high-energyphosphate pathway, and furthermore, showed evidence of antagonism ofcyclosporine-induced reduction of rat-brain, high-energy phosphate. Becauseinhibition of high-energy phosphate metabolic pathways has been postulated asa potential mechanism of cyclosporine-related neurotoxicity andnephrotoxicity, these unique properties of RAD may have potential clinicalbenefits (15).
RAD,in combination with Neoral® and corticosteroids, was effective in reducingthe severity of acute rejection episodes, particularly at doses of 2 mg or 4mg/day, suggesting that RAD may prove to be a useful addition to theimmunosuppressive armamentarium in clinical transplantation. Large-scale,controlled studies are currently in progress to further delineate the benefitsof this promising immunosuppressiveagent.
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