Hironori Takeuchi

Clinical significance of glucocorticoid pharmacodynamics assessed by antilymphocyte action in kidney transplantation : marked difference between prednisolone and methylprednisolone

A number of studies have demonstrated the impact of glucocorticoid response of peripheral lymphocytes on kidney allograft survival, suggesting that the better the glucocorticoid selection, the better the clinical outcome. However, individual differences in pharmacodynamics of clinically important glucocorticoids have not been taken into account. Four glucocorticoids (hydrocortisone, prednisolone, methylprednisolone, and dexamethasone) were examined for their ability to suppress in vitro blastogenesis of mitogen-stimulated PBL obtained from 122 chronic renal failure (CRF) patients waiting for renal transplantation and 98 healthy volunteers. Concentrations of steroids that gave 50% inhibition of lymphocyte blastogenesis (IC50) were determined individually in order to compare steroids and subject groups. Graft outcomes in 36 kidney transplant recipients treated with prednisolone were compared retrospectively with the prednisolone pretransplant IC50 values. Lymphocyte response to each glucocorticoid showed wide deviations among the subjects. Prednisolone IC50 values of the CRF patients showed the largest deviation, ranging from 1.0 to 10,000 micrograms/L. Thus, a significantly large population of the CRF patients (26.2%), when compared with the healthy subjects (4.1%) showed a marked decrease in lymphocyte response to prednisolone (P < 0.01). The binding capacity and affinity of lymphocyte glucocorticoid receptors did not differ significantly between the responders and nonresponders, suggesting that steroid resistance is a post-receptor event. The antilymphocyte potency of prednisolone assessed by IC50 of the steroid was less than that of hydrocortisone, whereas methylprednisolone was > 12-fold superior to prednisolone. After kidney transplantation, CRF patients who showed impaired preoperative lymphocyte response to prednisolone had a significantly high incidence of acute allograft rejection under prednisolone/CsA therapy (P < 0.01). It is concluded from these results that methylprednisolone could be of benefit to prednisolone-resistant recipients, who can be identified by the preoperative lymphocyte culture.

Immunosuppressant pharmacodynamics on lymphocytes from healthy subjects and patients with chronic renal failure, nephrosis, and psoriasis: Possible implications for individual therapeutic efficacy

In organ transplantation, patients with peripheral blood mononuclear cells (PBMCs) that exhibit resistance to cyclosporine (INN, ciclosporin) or glucocorticoids in vitro are refractory to therapy based on these drugs in vivo. However, detection or distribution of the resistant patients with immunologic disorders remains to be documented.

Glucocorticoids and cyclosporine induce apoptosis in mitogen-activated human peripheral mononuclear cells

Induction of apoptosis by immunosuppressive agents such as glucocorticoids (GCs) and cyclosporine (CsA) in cultured lymphoid cells has been suggested. However, there are few studies which demonstrate the induction of apoptosis by these agents in the activation process of human peripheral blood mononuclear cells (PBMCs). Here we show that potent immunosuppressive GCs and CsA induce apoptosis in concanavalin A (con A)-activated human PBMCs. In this study, GCs and CsA suppressed human PBMC-blastogenesis when activated by con A in a dose-dependent manner, where healthy PBMCs treated with > 100 ng/ml of each immunosuppressive agent exhibited a DNA-ladder structure in electrophoretic analysis. In three chronic renal failure (CRF) patients, dose-dependency of the PBMC-apoptosis induction was confirmed by our quantification of fragmented DNA using ELISA. Furthermore, the enrichment of DNA fragmentation was significantly associated with the rate of PBMC-blastogenesis when treated with GCs or CsA (r = -0.466, P < 0.01). These results suggested that suppression of the mitogen-induced PBMC-blastogenesis by the immunosuppressive agents should be correlated with the induction of apoptosis.

Safety and efficacy of conversion from twice-daily tacrolimus (prograf) to once-daily prolonged-release tacrolimus (graceptor) in stable kidney transplant recipients.

BACKGROUND Graceptor is a new modified-release once-daily formulation of tacrolimus with an efficacy and safety profile similar to twice-daily tacrolimus (Prograf), as identified by clinical trials, offering a more convenient dosing regimen to improve adherence. The aim of this study was to analyze the safety of a 1:1 dose conversion from twice-daily Prograf to once-daily Graceptor in stable kidney transplant recipients. METHODS We switched 33 Japanese patients who had undergone kidney transplantation ≥1 years before from twice-daily Prograf to once-daily Graceptor. The dose conversion ratio between Prograf and Graceptor was 1:1. We compared the following parameters: minimum tacrolimus concentration (C(min)); concentration dose per weight (CDW); serum creatinine (sCr); blood urea nitrogen (BUN); total cholesterol (TC); high-density lipoprotein cholesterol (HDL-C); uric acid (UA); fasting blood sugar (FBS). Time points for measurements were 1 month before study start and 1 and 2 months afterward. RESULTS The mean age of the subjects in this study was 46.5 ± 13.1 years. Mean C(min) decreased from 4.55 ± 1.79 to 3.20 ± 1.22 ng/dL. The mean CDW also decreased, from 99.8 ± 69.5 to 75.0 ± 55.1 mg/dL/kg over the 2 months. There were no significant changes in sCR, BUN, UA, and FBS. Mean TC increased from 187.5 ± 51.4 to 194.3 ± 43.4 mg/dL, and mean HDL-C changed from 53.7 ± 12.0 to 56.1 ± 11 mg/dL. There were no episodes of rejection or infection. CONCLUSIONS We conclude that switching from Prograf to Graceptor is safe and has the advantage of improving adherence. It could also have a beneficial effect in controlling glycemic levels and the adverse effects of tacrolimus. In many cases (25%-30%), the minimum concentration of tacrolimus decreased after changing tablets. With Graceptor, the ratio of area under trough level to area under the curve (AUC) is low compared with Prograf, resulting in low C(min) values of 1-2 ng/mL, and the AUC for Graceptor is very similar to that for Prograf.

Comparative study of the cellular pharmacodynamics of calcineurin inhibitors between patients with chronic renal failure awaiting renal transplantation and cirrhosis patients awaiting liver transplantation.

The in vitro response of peripheral blood mononuclear cells (PBMCs) to the suppressive effects of calcineurin inhibitors is known to correlate with the clinical efficacy of drugs used in renal transplantations. The present study was conducted to examine the differences of PBMC responses to calcineurin inhibitors between chronic renal failure (CRF) patients awaiting renal transplantation and cirrhosis patients awaiting liver transplantation. The study included 99 CRF patients awaiting renal transplantation and 27 cirrhosis patients awaiting liver transplantation. Twenty milliliters of venous blood was taken 1–7 days before transplantation. The in vitro drug concentrations giving 50% inhibition of PBMC blastogenesis stimulated with concanavalin A (IC50s) were calculated. The suppressive effects of tacrolimus against PBMC blastogenesis were more than 10–100 times stronger than those of cyclosporine. The median IC50 value for cyclosporine against the CRF PBMCs was not significantly different from the median IC50 value against the cirrhosis PBMCs. In contrast, tacrolimus sensitivity in cirrhosis PBMCs is approximately seven times higher than that in CRF PBMCs. The median IC50 value for tacrolimus against cirrhosis PBMCs was significantly lower and therefore the effect was stronger in comparison to the CRF PBMCs (p < 0.001). These data suggest that the PBMCs of cirrhosis patients, in comparison to those of CRF patients, are highly sensitive to the suppressive effect of tacrolimus. However, PBMC sensitivity to cyclosporine was not significantly different between the CRF and cirrhosis patients. These observations raise the possibility that treatment with tacrolimus, rather than cyclosporine, may therefore be a better choice to reduce the risks of allograft rejection in liver transplantation.

A comparison of prednisolone and methylprednisolone for renal transplantation.

A large difference in immunosuppressive potency between methylprednisolone and prednisolone has been suggested in vitro. However, the selection of the best glucocorticoid for renal transplantation has been seldom considered so far. Thus, the present study was undertaken to compare therapeutic efficacy between prednisolone and methylprednisolone in renal transplantation. We studied 42 renal transplant recipients who were operated on between 1990 and 1994. The patients were divided into two treatment groups: a methylprednisolone/cyclosporine group (n=19) and a prednisolone/cyclosporine group (n=23). Clinical outcome and drug side effects were compared retrospectively between the treatment groups 24–84 months after transplantation. The overall graft survival time in patients treated with methylprednisolone/cyclosporine was superior to that in patients treated with prednisolone/cyclosporine (p<0.05). Among the recipients from cadaver donors, 5/16 (31.3%) treated with prednisolone required nephrectomy, whereas none of the 10 patients treated with methylprednisolone received nephrectomy (p<0.01). An examination of the recipients from living related donors revealed that serum creatinine levels 24–36 months after operation were significantly lower in the methylprednisolone group (p<0.05). Cyclosporine trough levels and glucocorticoid side effects were similar between the treatment groups. The results raised the possibility that methylprednisolone is superior to prednisolone when combined with cyclosporine for maintenance immunosuppressive therapy in renal transplantation.

Glucocorticoid-resistance in peripheral-blood lymphocytes does not correlate with number or affinity of glucocorticoid-receptors in chronic renal failure patients

Glucocorticoid (GC) resistance in patients with chronic renal failure (CRF) seriously impairs successive GC therapy after renal transplantation. We examined the relationship between GC-receptor (GC-R) parameters in peripheral-blood mononuclear cells (PBMC) and PBMC resistance to GC in 21 CRF patients and 18 healthy subjects. Each subject group was divided into two subgroups according to PBMC sensitivity to prednisolone in a mitogen assay procedure; i.e., sensitive (IC50 < 381 ng/mL) and resistant (IC50 > 381 ng/mL) groups. In healthy subjects, the mean GC-R Bmax and Kd in quiescent PBMC of the GC-sensitive group were 2.89 +/- 1.23 fmol/10(6) cells and 4.00 +/- 2.24 nM, respectively. The Bmax in these subjects significantly increased to 6.61 +/- 2.02 (257.7 +/- 107.8%) after 24 h stimulation with concanavalin A (p < 0.01), while the Kd change was not significant. The GC-R Bmax and Kd in quiescent PBMC of the GC-resistant group were 5.33 +/- 1.37 fmol/10(6) cells and 3.20 +/- 1.39 nM, respectively. Both of these parameters, however, did not change significantly after mitogen stimulation. There was a significant negative correlation between IC50S of prednisolone and increase-ratios (post/pre ratio) of Bmax after mitogen stimulation (p < 0.05). In CRF patients, Bmax and Kd in quiescent PBMC of the GC-sensitive group were 6.04 +/- 2.35 fmol/10(6) cells and 3.49 +/- 1.72 nM, respectively, while those in PBMC of the GC-resistant group were 5.13 +/- 2.31 fmol/10(6) cells and 4.04 +/- 1.62 nM, respectively. The Bmax and Kd were not significantly changed after mitogen stimulation in both subgroups of CRF. Moreover, in contrast to healthy subjects, there was no correlation between IC50 and GC-R parameters in CRF. We concluded that, in healthy subjects, decreased PBMC capacity to amplify GC-R numbers in response to mitogen is correlated with GC resistance, whereas in CRF patients the resistant mechanism is not correlated with GC-R parameters. An unknown event might be involved in GC-resistance of CRF.

Early Steroid Withdrawal in Adult Kidney Transplantation at a Single Center

BACKGROUND Beneficial effects of protocols using minimal steroid exposure have been recently reported. The purpose of this study was to evaluate the outcomes of kidney transplantation recipients who received immunosuppression protocols with early steroid withdrawal (ESW) at our center. METHODS We retrospectively studied 84 kidney transplant recipients who had received ESW immunosuppressive protocols at our center from March 2005 to December 2010. The immunosuppressive regimen was a combination of calcineurin inhibitors (tacrolimus/cyclosporine), methylprednisolone, which was tapered and discontinued within 2 months, mycophenolate mofetil, and basiliximab (postoperative days 0 and 4). We compared the outcomes of our ESW recipients with those of a historical control group (February 2003 to January 2005; n = 18). RESULTS Clinical acute rejection episodes were observed in 15 (17.9%) and 5 (27.8%) cases in the ESW and control groups, respectively. Cytomegalovirus infection occurred in 12 (14.3%) and 5 (27.8%) cases in the ESW and control groups, respectively. The creatinine levels at 1 year after transplantation were 1.3 ± 0.4 mg/dL and 1.3 ± 0.5 mg/dL in the ESW and control groups, respectively. In the ESW group of 84 recipients, actuarial patient survival at 1 year was 94.0%. In the historical group of 18 recipients, the actuarial patient survival at 1 year was 100% (P = .76). In the ESW group the graft survival rate at 1 year was 95.2%. In the historical group, graft survival rate at 1 year was 100% (P = .65). There were no significant differences in the parameters between the groups. CONCLUSIONS The outcomes from this study were considered to be acceptable; however, the possibility of improving the protocols exists.

Falsely Abnormally Elevated Blood Trough Concentration of Tacrolimus Measured by Antibody-Conjugated Magnetic Immunoassay in a Renal Transplant Recipient: A Case Report

This report presents a falsely abnormally elevated blood trough concentration (C(t)) of tacrolimus measured by antibody-conjugated magnetic immunoassay (ACMIA) methods in a renal transplant recipient. Because the C(t) of tacrolimus was 78.5 ng/mL at day 2 after a 52-year-old man underwent renal transplantation, we stopped the tacrolimus extended-release formulation. However, because the abnormally elevated blood C(t) continued in the range of 41.1-59.1 ng/mL, we then measured the tacrolimus concentration in a stored blood sample before renal transplantation, it was 43 ng/mL. Consequently, the day-7 blood sample was measured with both ACMIA and enzyme-linked immunoassay, showing C(t) values of 42.8 ng/mL and 0.89 ng/mL, respectively. Because the abnormally elevated C(t) was falsely measured by the ACMIA method, we restarted tacrolimus However, the calcineurin inhibitor was subsequently converted to cyclosporine at day 21 after renal transplantation. Although cyclosporine was also measured by ACMIA, there was not an abnormally elevated C(t). Subsequently, the tacrolimus concentration ratio in plasma and whole blood (P/B-tacrolimus concentration ratio) was measured by ACMIA in a posttacrolimus blood sample. The P/B-tacrolimus concentration ratio was 100%. In contrast, the P/B-tacrolimus concentration ratio was <30% in 2 control patients administered tacrolimus. It has been reported recently that there were cases showing falsely slightly elevated C(t) of tacrolimus within the therapeutic range of concentrations. Therefore, we must be careful not to reduce the tacrolimus dose falsely. We consider confirmatory methods for a falsely abnormally elevated C(t) of tacrolimus measured by ACMIA to (1) measure P/B-tacrolimus concentration ratio, (2) compare ACMIA with another measurement, and (3) evaluate a blood sample stored before tacrolimus administration.

Clinical significance of the cellular pharmacodynamics of tacrolimus in living-donor liver transplantation.

Successful immunosuppressive therapy is critical for liver transplantation; however, a considerable number of patients experience fatal rejection or alternatively exhibit serious infection resulting from excessive immunosuppression. The in vitro tacrolimus response of peripheral blood mononuclear cells (PBMCs) before transplantation was compared to the clinical outcome up to 4 weeks after operation in 28 living-donor liver transplant recipients treated with tacrolimus. The tacrolimus IC50 values against concanavalin A-induced PBMC blastogenesis in vitro were calculated. These recipients were classified into two groups with the mean tacrolimus IC50 (0.18 ng/ml) as the cutoff point, after which the clinical outcome between the patient groups was compared. The allograft rejection incidence in the low-sensitivity group (IC50 < 0.18 ng/ml; n = 16) was 6/12 (50.0%), which was significantly higher than the incidence of 2/16 (12.5%) in the high-sensitivity group (IC50 > 0.18 ng/ml; n = 12) (p = 0.0297). In contrast, the infection incidence in the high-sensitivity group was 6/16 (37.5%), which was significantly higher than that of the low-sensitivity group (1/12; 8.3%) (p = 0.0401). These data suggest that patients exhibiting a low PBMC sensitivity to tacrolimus have a risk of rejection, whereas highly sensitive patients have a risk of infection in living-donor liver transplantations under tacrolimus therapy.