Calvin Stiller

A randomized, prospective multicenter pharmacoepidemiologic study of cyclosporine microemulsion in stable renal graft recipients. Report of the Canadian Neoral Renal Transplantation Study Group.

BACKGROUND The safety, tolerability, and pharmacokinetics of conventional cyclosporine (ConCsA) and cyclosporine microemulsion (MeCsA) were compared under conditions of normal clinical practice in a prospective, randomized, concentration-controlled, pharmacoepidemiologic study. METHODS Between September 1994 and March 1995, 1097 stable renal transplant recipients in 14 Canadian centers were randomized 2:1 to treatment with MeCsA or ConCsA. Patients were commenced on each study drug at a dose equal to their previous therapy with ConCsA, and the dose was adjusted to maintain predose whole blood cyclosporine concentrations within the therapeutic range established for each center. Prednisone and azathioprine were continued unless dose adjustment was required for clinical reasons. RESULTS The mean cyclosporine concentration was comparable in both treatment groups at all time points throughout the 6 months of follow-up. The mean dose of cyclosporine was 3.6 mg/kg/day in both treatment groups at entry to the study, and declined by 0.3% and by 2.8% in patients receiving ConCsA and MeCsA, respectively. The nature and severity of adverse events were similar in both treatment groups, but there was a transient increase in neurological and gastrointestinal complications in the group receiving MeCsA within the first month after conversion (P<0.05). Serum creatinine and creatinine clearance did not change in either treatment group throughout the study. Biopsy-proven acute rejection occurred in three patients (0.8%) receiving ConCsA and in seven patients (0.9%) receiving MeCsA, with non-histologically proven acute rejection in an additional three patients (0.8%) receiving ConCsA and five patients (0.6%) receiving MeCsA (P=NS). Serum creatinine rose transiently in 35 patients (9.8%) receiving ConCsA and 138 patients (18.7%) receiving MeCsA (P<0.05) and resolved either spontaneously or after a reduction in the cyclosporine dose. One graft was lost in the MeCsA group due to irreversible rejection, and seven patients died, three in the group receiving ConCsA and four of those receiving MeCsA (P=NS). Absorption of cyclosporine was more rapid and complete from MeCsA than from ConCsA during the first 4 hr of the dosing interval, resulting in almost 40% greater exposure to the drug (P<0.001). There was close correlation between area under the time-concentration curve (AUC) over the first 4 hr of the 12-hr dosage interval and AUC over the entire 12-hr dosage interval for both formulations, making AUC over the first 4 hr a good predictor of total cyclosporine exposure. Using this parameter, patients with low absorption randomized to receive MeCsA showed a marked increase in drug exposure by months 3 and 6, whereas there was no change in those who continued on ConCsA. A limited sampling strategy utilizing samples at the predose and postdose trough levels provided an excellent correlation with drug exposure, particularly for patients receiving MeCsA (r2=0.94 MeCsA vs. r2=0.89 ConCsA). CONCLUSIONS MeCsA appears to be a safe and effective therapy in stable renal transplant patients and provides superior and more consistent absorption of cyclosporine when compared with ConCsA. Transient toxicity after conversion to MeCsA occurs in some patients, and may reflect the increased exposure to cyclosporine. Use of a limited sampling approach combining trough and 2-hr postdose concentrations may provide an effective way to monitor this exposure.

Pattern of liver, kidney, heart, and intestine allograft rejection in different mouse strain combinations.

With advances in microsurgery and molecular biology, the mouse model for organ transplantation has become increasingly popular. However, knowledge about these models is limited, as only a small number of centers have experience with murine models. In this study, we compared the rejection pattern after liver, kidney, heart, and small bowel transplantation in the three different mouse strain combinations: (1) C57BL/6 (H2b)-->BALB/c (H2d), (2) BALB/c (H2d)-->CBA (H2k), and (3) C57BL/6-->C3H/HeN (H2k). Our study demonstrated that mouse allograft survival varies depending on the organ graft and on the donor-recipient strain combinations. The majority of liver allografts were spontaneously accepted despite complete MHC disparity. A mixed pattern of acute rejection and acceptance occurred in kidney recipients depending on the donor-recipient strain combination. All the heart grafts developed rejection and all the intestinal grafts were rapidly rejected with no spontaneous acceptance. The criteria for rejection, the potential applications, and the limitations of each model are discussed. The models described in this article provide a number of useful choices for organ transplantation research.

Intestinal permeability and bacterial translocation following small bowel transplantation in the rat.

In addition to its role in absorbing nutrients, the intestinal mucosa provides an important barrier against toxins and bacteria in the bowel lumen. The present study evaluated gut barrier function following orthotopic (in continuity) intestinal grafting in rats. Graft histology, intestinal permeability, and bacterial translocation to the grafted mesenteric lymph nodes, the hosts liver, and the hosts spleen were assessed on the 3rd, 5th, and 7th postoperative days. The study group received no immunosuppression after allotransplantation. The two control groups included rats with isografts and rats with cyclosporine-treated allografts. On the 7th POD, the study animals had moderate transmural inflammation due to rejection, with normal histology in the isografts and CsA-treated allografts; increased intestinal permeability, measured by urinary excretion of oral 51Cr-EDTA (P less than 0.01); and increased number of bacteria in the MLN and spleen (P less than 0.05). The number of bacteria in the MLN and spleen of the study group positively correlated with the changes in intestinal permeability (P less than 0.05). Rejection of the orthotopic intestinal graft leads to increased intestinal permeability and bacterial translocation from the lumen of the graft to the hosts reticuloendothelial system. Measures to improve gut barrier function and antibiotic therapy during rejection episodes may help reduce the incidence of septic complications after intestinal grafting.

Safety and tolerability of cyclosporine and cyclosporine microemulsion during 18 months of follow-up in stable renal transplant recipients: a report of the Canadian Neoral Renal Study Group.

BACKGROUND There has been concern that the increased drug exposure associated with treatment with cyclosporine microemulsion (CsA-ME) would lead to an increase in adverse events. METHODS The long-term safety and tolerability of conventional cyclosporine (CsA) and CsA-ME were compared in a randomized, multicenter, pharmacoepidemiologic study involving 1097 stable renal transplant patients after 18 months of follow-up. RESULTS No significant difference was seen in change in serum creatinine or calculated creatinine clearance between the two groups. Episodes of deterioration in renal function (change in serum creatinine > or = 20%) were categorized with the following results for CsA-ME versus CsA, respectively: acute rejection, 4.5% vs. 4.5%; chronic rejection, 8% vs. 11%; CsA nephrotoxicity, 12% vs. 7% (P=0.008); transient changes, 17% vs. 12%; other causes, 4% vs. 6%. During the first 6 months of the study, a transient increase in the incidence of gastrointestinal and neurological adverse events was seen in the CsA-ME group compared with the CsA group. Up to 18 months, patients in the CsA group reported significantly fewer hearing and vestibular disorders, but more cardiovascular problems than those in the CsA-ME group (P=0.035). CONCLUSIONS Tolerance to CsA and CsA-ME was similar. Renal function over 18 months was not adversely affected by the increased drug exposure with CsA-ME, although there was a transient increase in nephrotoxicity. The frequency of acute and chronic rejection did not change.

Histological and immunopathological analysis of recovered encapsulated allogeneic islets from transplanted diabetic BB/W rats

Allogeneic islets encapsulated in an alginate/poly-L-lysine membrane and transplanted into diabetic BB/W rats resulted in graft failure within 2 weeks of transplantation. Graft failure was associated with a dense pericapsular infiltrate (PCI) that resulted in necrosis of the encapsulated islets. The PCI could be inhibited by immunosuppressive agents, including cyclosporine and dexamethasone, and this resulted in a significant increase in graft survival. Immunopathological characterization of the PCI indicated that there was a predominance of macrophages. T helper cells also appeared to be present in this PCI. Empty capsules were also found to induce a similar PCI that was identical in composition to that found around encapsulated islets. Thus alginate/poly-L-lysine capsules do not appear to be biocompatible and may account for the variable results in islet graft survival found with these capsules.

Effect of cyclosporine on urinary prostanoid excretion, renal blood flow, and glomerulotubular function.

The clinical usefulness of cyclosporine (CsA) in organ transplantation and autoimmune diseases is limited by its intrinsic nephrotoxicity. The mechanism of this renal impairment was examined utilizing an animal model in which male Sprague-Dawley rats were administered oral CsA in doses of 25, 37.5, and 50 mg/kg/day for 7 days and 50 mg/kg/day for 2, 4, and 7 days. Urinary thromboxane B2 (TXB2) excretion increased from 30.6± 2.3 to 60.8±4.4 ng/24 hr I<0.001, following 48 hr of CsA dosing. In addition, a concomitant rise in proximal tubular sodium reabsorption was observed with fractional excretion of sodium decreasing from 0.502±0.091 to 0.223 ± 0.037% P<0.05. Urinary prostaglandin E2 and 6-keto-prostaglandin F1α excretion increased twofold, although plasma levels of all 3 prostanoids did not vary from cpntrols. Functional changes included decreases in the relative renal blood flow of 53% P<0.05, and the clearance of creatinine and urea of 46% and 42%, respectively on day 7 of treatment, while renal

Successful intestinal transplantation in pigs treated with cyclosporine.

To date, it has not been possible to reliably prevent intestinal allograft rejection in large animals. This study was undertaken to determine if continuous i.v. cyclosporine (CsA) followed by p.o. CsA would prevent rejection in outbred piglets with orthotopic, in-continuity intestinal allografts. Untreated recipients (n=7) died of rejection (2), interstitial pnuemontitis (3), or technical complications (2) at 5.3±1 days. Intestinal recipients treated with i.v. CsA 8 mg/kg/day and i.v. steroids (n=3) died of rejection (mean survival 11.3±3.2 days). CsA 20 mg/kg/day i.v. plus i.v. steroids for 21 days, followed by p.o. CsA 25 mg/kg/day (n=6) prevented rejection; however, most of the recipients developed fatal infections (mean survival 28±8 days). Intravenous CsA 15 mg/kg/day for 7–10 days (n=16), followed by p.o. CsA 30 mg/ kg/day in tapering doses reliably prevented graft rejection, permitting long-term survival (mean survival 121±32 days). Rejection did not occur in 7 animals when CsA was discontinued at 97±11 days. Seven animals surviving more than 100 days maintained normal nutritional indices and gained weight at the same rate as control animals. This study provides a rationale for further experimentation to determine the feasibility of intestinal transplantation in man.

A comparison of heterotopic and orthotopic intestinal transplantation in rats.

Two surgical techniques are commonly used for small intestinal transplantation: heterotopic (accessory) intestinal grafting (HIT), where the small bowel is initially defunctioned with restoration of intestinal continuity at a later date, and orthotopic (in continuity) intestinal grafting (OIT), where the small bowel is immediately anastomosed to the native intestine. The present experiments were undertaken to compare the advantages and disadvantages of these two surgical models. Graft barrier function (intestinal permeability), intestinal histology, and graft survival were evaluated after heterotopic and orthotopic intestinal transplantation in the following groups of rats: group 1: isografts, group 2: untreated allografts, group 3: low-dose cyclosporine—treated allografts (subcutaneous CsA 2 mg/kg/day), and group 4: high-dose CsA-treated allografts (subcutaneous CsA 4 mg/kg/day). Intestinal permeability was consistently higher after HIT than OIT in all of the groups (ANOVA; P<0.01). Histological evidence of rejection appeared earlier after HIT than OIT (HIT 5th postoperative day (POD); OIT 7th POD; P<0.05). The mean survival of untreated allografts was longer after HIT than OIT (HIT 15.7±6 days, OIT 9.2±1 days, P<0.05). The rats treated with low-dose CsA after OIT lost weight and died of rejection after a mean survival time of 17.7±2 days, while the rats treated with low-dose CsA after HIT remained well until sacrifice on POD 28 (P<0.01). The rats with isografts and rats with allografts treated with high-dose CsA remained well after HIT and OIT until sacrifice on the 28th POD. These data suggest that nutrients and other factors in the succus entericus may improve gut barrier function and delay the onset of rejection after OIT. However, rejection of the orthotopic intestinal graft is usually fatal, while rejection of the heterotopic graft is often surprisingly well tolerated. These factors must be taken

Transplantation of encapsulated allogeneic islets into diabetic BB/W rats : effects of immunosuppression

Allogeneic islets obtained from Lewis rats were transplanted into diabetic BB/W rats with or without cyclosporine. In addition, these islets were encapsulated in alginate-poly L-lysine membranes and then transplanted into diabetic BB/W rats with or without immunosuppressive and/or antiinflammatory agents. The agents used were cyclosporine, dexamethasone, indomethacin (Ind), or a combination of these. Our results show that islets alone survived for 7 days, with or without CsA therapy. Encapsulated islets survived for 14.2 days, and this was extended by CsA, Dex, or CsA + Ind. Loss of encapsulated graft functions was associated with formation of a dense pericapsular infiltrate, which was inhibited by CsA, Dex, CsA + Ind, or CsA + Dex. In addition, the infiltrate was reduced in animals that had diabetes for long periods of time (greater than 5 months versus less than 1 month). Empty capsules also provoked this cellular response. Thus, encapsulation of islets resulted in slightly prolonged islet survival, which was further enhanced by immunosuppression.

Graft-versus-host disease associated with intestinal transplantation in the rat. Host immune function and general histology.

Graft-versus-host disease was studied on the 10th and 14th postoperative days in Lewis x Brown Norway F1 rats (LBN-F1) receiving Lewis accessory heterotopic intestinal allografts. LBN-F1 isograft recipients and LBN-F1 rats were used as controls. The rats were injected with sheep erythrocytes five days before sacrifice. Rats with graft-versus-host disease had progressive loss of the normal architecture of the lymphoid organs. Skin, liver, colon, and salivary glands were infiltrated with immunoblasts and had patchy areas of necrosis. Concurrent with these changes, there were significant, progressive reductions in hemolytic titers, splenocyte plaque-forming counts, viable splenocytes, and the in-vitro splenocyte response to stimulation with concanavalin A. Graft-versus-host disease following intestinal allotransplantation damages the hosts lymphoid tissues, producing profound immunosuppression. This finding has implications for clinical intestinal transplantation.