Efficacy and safety of low-dose everolimus and tacrolimus in kidney transplant recipients: A retrospective study

Abstract

BACKGROUND

Standard immunosuppressive regimens result in low rejection rates in the first year after transplantation. However, long-term survival rates after renal transplantation remain relatively poor and are associated with drug-induced side effects.

AIM

To assess the effects of switching from traditional mycophenolate mofetil (MMF) combined with steroids and tacrolimus to a low-dose regimen of everolimus and tacrolimus in post-kidney transplant patients, with the aim of reducing the adverse effects of MMF and high-dose calcineurin inhibitors.

METHODS

This was a retrospective, single-center study of patients who received a combination therapy of steroids + tacrolimus (low dose) + everolimus (low dose) after kidney transplantation (KT). Data was obtained from patients who underwent KT between 2007 and 2019. Everolimus was initiated at 0.25 mg twice daily, aiming for pre-dose concentrations of 5-7 ng/mL (combined trough levels of tacrolimus and everolimus). Improvement in graft survival was assessed by measuring serum creatinine, urinary protein, and lipid profiles. Safety studies were conducted to observe drug-induced side effects.

RESULTS

We included 36 patients from a single center. After switching to everolimus, there was a significant reduction in serum creatinine levels which remained stable over one year. Cholesterol levels also improved and remained stable in most patients. Half of the patients reported improvement in their cholesterol levels and continuous stability. Vomiting (47.1%) and diarrhea (32.4%) were the most common adverse effects of MMF treatment. Other side effects included urinary tract infections (20.6%) and anemia (5.9%).

CONCLUSION

Combination therapy with everolimus (low dose) + tacrolimus (low dose) was superior to MMF + tacrolimus in terms of a lower incidence of side effects and stable serum creatinine levels over long-term follow-up.

Key Words: Mammalian target of rapamycin inhibitors; Everolimus; Calcineurin inhibitors; Immunosuppression; Graft rejection; Kidney transplant

Core Tip: The use of combination of low dose tacrolimus and everolimus is not common in kidney transplant recipients. We show in our study that the combination therapy with everolimus (low dose) + tacrolimus (Tac) (low dose) as compared to mycophenolate mofetil + Tac is found to be superior in terms of lower incidence of side effects and stable serum creatinine levels over long-term follow-up.

INTRODUCTION

Kidney transplantation (KT) has evolved into the treatment of choice for patients with acute and chronic end-stage kidney disease. However, it is associated with key concerns, such as acute kidney graft rejection and the preservation of long-term kidney function[1,2]. To overcome these issues and improve patient outcomes, immunosuppressants (IS) are administered in various combinations before, during, and after KT[3]. The most commonly used treatment regimens include administering a distinct IS initially for induction and later for maintenance. Either interleukin-2 receptor antibodies or anti-thymocyte globulin is used during and immediately post KT, while a combination of several IS is used for maintenance therapy. Frequently used maintenance drugs include glucocorticoids, antimetabolites, calcineurin inhibitors (CNIs), mammalian target of rapamycin inhibitors (mTORi), and the co-stimulation blocker belatacept. The combined administration of induction and maintenance immunosuppressive therapy is considered superior to maintenance therapy alone, with evidence of improved graft survival[3-6].

Most clinical studies have focused on graft function and graft rejection during the first year post-transplantation. However, very few studies have evaluated the effects of drugs in various combinations during the maintenance phase, with the goal of reducing the associated side effects[3,6-9].

Tacrolimus (Tac), a commonly used CNI, causes side effects such as headache, diarrhea, dyspepsia, vomiting, and tremors, whereas cyclosporine A frequently induces hypertension, gingival hyperplasia, and hirsutism. CNI-induced nephrotoxicity due to its prolonged exposure remains the most poorly tolerated adverse effect by patients[10-12]. The Kidney Disease-Improving Global Outcomes guidelines recommend a triple combination of CNIs, corticosteroids, and antimetabolites[13]. Mycophenolate, a first-line antimetabolite available as mycophenolate mofetil (MMF) or enteric-coated mycophenolate sodium, causes gastrointestinal (GI) side effects and other complications, including viral infections[14,15]. An alternative regimen has been suggested in certain clinical studies, advocating the addition of everolimus (Evr) instead of MMF to Tac to reduce CNI-related nephrotoxicity[16-18].

Unfortunately, few follow-up reports are available on the prolonged concomitant treatment of Evr with low-dose Tac in kidney transplant patients. Therefore, long-term follow-up studies are needed to evaluate the efficacy of the combination treatment of Evr and Tac in de novo kidney transplant recipients. The current study reviews the long-term benefits of combining low-dose Evr and low-dose Tac, as well as the associated improvements in the side-effect profile in post-kidney transplant patients.

MATERIALS AND METHODS

Study design

This was a retrospective, single-arm, single-center study conducted at Medica Hospital, Kolkata. The data were obtained from the medical records of patients who had undergone KT and were initially treated with MMF + steroid + CNI (Tac) post-transplantation. Data was obtained from patients who underwent transplantation between 2007 and 2019.

Study objective

The objective of this study was to evaluate the effectiveness of the combination therapy [steroid + Tac (low dose) + Evr (low dose)] in reducing MMF-associated side effects while preserving kidney function in post-kidney transplant patients. This study recorded and evaluated changes in certain parameters in post-kidney transplant patients who were initially on the MMF regimen [MMF + steroid + CNI (Tac)] and later switched to the combination protocol [steroid + Tac (low dose) + Evr (low dose)]. The parameters considered were as follows: Baseline serum creatinine levels were measured before and after initiating combination therapy [steroid + Tac (low dose) + Evr (low dose)] to assess renal function; Proteinuria and lipid profile; Maintenance of combined trough levels of 5-7 ng/mL for both drugs (Tac level 3-4 ng/mL; Evr 2-3 ng/mL); Side effects reported by the patients both before and after initiating the combination therapy; The dosage of Tac and Evr administered to patients was 0.25 mg twice daily. The trough levels for both drugs were defined as 5-7 ng/mL. Levels were measured using liquid chromatography/mass spectrometry in a single laboratory at scheduled intervals.

Study population and setting

The study population consisted of 70 post-renal transplant patients of either sex who were experiencing MMF or its sodium salt-associated side effects during maintenance therapy and were subsequently switched to combination therapy with reduced doses of Tac and Evr. However, only 36 patients were included in the study based on the inclusion and exclusion criteria. Of the 34 patients who were excluded from this study, 15 were foreign patients in which local physicians changed the doses by themselves, 14 had grossly inadequate follow-up, 2 died during coronavirus disease with functioning graft and the remaining 3 switched back to MMF due to unavailability or higher cost of everolimus.

Patients who presented with vomiting for 3 consecutive months or more than three episodes over a 3-month period, and three or more urinary tract infections (UTI) in the 6-month period were switched to combination therapy. However, all causes of vomiting due to upper GI disturbances were excluded before prescribing MMF or its salts as the offending agents. Similarly, anatomical causes of UTI were ruled out before presuming MMF to be the causative agent.

Urine protein levels and lipid profiles were determined before and after the initiation of combination therapy. Patients with proteinuria > 800 mg/day or hyperlipidemia were not administered the combination treatment.

The transition from MMF to Evr was performed with a 2-week overlap. Upon reporting any evidence of rejection, the treatment regimen was switched back to the MMF/salt protocol, in addition to an increased steroid dose for the next month. Patients receiving the combination treatment of steroids + Tac (low dose) + Evr (low dose)] were closely monitored for proteinuria, edema, and hyperlipidemia. Patients with a urine protein level of over 1 g/day were switched back to the MMF/salt protocol. Patients with increased hyperlipidemia were switched back to the MMF protocol. Additionally, a decline in urine protein and blood lipid levels was considered a sign of improved graft survival.

Patients’ inclusion and exclusion criteria

Patients who reported intolerance to MMF or sodium salts were included. International patients with incomplete data or follow-up, those with a follow-up period, and those who died during treatment were excluded from the study. Patients who developed graft rejection or significant kidney disease during the maintenance phase, or who had to switch back to MMF, were excluded from the final data analysis. Patient data were analyzed from the initiation of combination therapy through the last dose received, as per the records.

Statistical analysis

Changes in serum creatinine levels before and after combination therapy were compared using paired t-tests. The average trough values for both drugs were compared using paired t-tests at 1 week, 6 months, and 1 year after transplantation. Quantitative and qualitative analyses of urine protein levels were performed using the paired t-test and χ² test, respectively. The level of significance was set at P < 0.05. Statistical analyses were conducted using SPSS software.

Safety and tolerability assessment

Incidence safety was ensured by registering adverse events.

RESULTS

Of the 70 patients recruited for the study, only 36 post-kidney transplant patients were included in the analysis (May 2007 to December 2019). Of the total study population, 72% (n = 26) were males. The demographic details are summarized in Table 1. Data from all 36 patients were included in the statistical analysis.

Table 1 Demographic data and timing of everolimus switch and creatinine results.

Patient ID	Age	Gender	Pre-transplant diabetes	Time from KT to switch (years)	Cr before switch (mg/dL)	Cr 1 month after (mg/dL)
KT-01	59	Male	Yes	2.5	1.13	1.22
KT-02	49	Male	No	2.3	1.30	1.18
KT-03	35	Male	No	2.8	1.27	0.98
KT-04	63	Male	Yes	1.9	0.78	1.21
KT-05	28	Female	No	1.2	1.59	1.17
KT-06	41	Male	No	2.4	1.57	1.76
KT-07	59	Male	Yes	2.5	1.45	1.02
KT-08	39	Male	Yes	2.1	1.34	1.98
KT-09	43	Female	Yes	2.5	0.80	1.07
KT-10	31	Male	Yes	0.8	1.26	1.25
KT-11	31	Female	Yes	2.0	1.29	1.70
KT-12	44	Male	Yes	1.9	1.08	0.90
KT-13	56	Male	No	1.1	1.38	1.47
KT-14	60	Male	Yes	1.2	1.64	1.89
KT-15	44	Female	Yes	1.1	2.32	0.75
KT-16	23	Female	No	2.3	1.53	1.45
KT-17	42	Male	Yes	1.6	1.57	1.48
KT-18	22	Male	No	2.0	1.42	1.68
KT-19	44	Male	Yes	2.8	0.60	0.90
KT-20	64	Male	No	1.5	1.44	0.87
KT-21	50	Male	No	1.8	1.48	1.58
KT-22	58	Female	Yes	2.5	2.58	1.50
KT-23	22	Male	Yes	1.5	1.36	1.48
KT-24	41	Male	No	1.2	1.59	1.52
KT-25	53	Male	No	1.6	1.43	1.11
KT-26	32	Female	No	1.3	0.91	1.47
KT-27	42	Male	No	2.9	1.98	1.49
KT-28	64	Female	Yes	2.6	1.80	1.10
KT-29	45	Female	Yes	0.8	1.81	2.11
KT-30	69	Male	Yes	2.7	1.03	1.56
KT-31	47	Male	No	2.6	2.10	0.92
KT-32	62	Female	No	1.4	0.81	1.64
KT-33	48	Male	Yes	2.8	1.72	1.40
KT-34	36	Male	Yes	2.1	2.46	1.69
KT-35	35	Male	Yes	2.6	0.99	2.03
KT-36	67	Male	Yes	2.8	1.19	1.06

KT: Kidney transplantation; Cr: Creatinine.

Renal function (serum creatinine)

The mean serum creatinine values before and after switching to Evr were 1.45 ± 0.46 mg/dL and 1.38 ± 0.39 mg/dL, respectively. After Evr administration, the mean value decreased by 5.2% (Table 2).

Table 2 Serum creatinine levels before and after initiation of everolimus.

Serum creatinine (mg/dL)	Mean	SD	P value
Before everolimus	1.44	0.46
After everolimus (post one month)	1.38	0.35	0.475
After everolimus (post one year)	1.41	0.44	0.398

P < 0.05.

Protein levels in urine and lipid profile

Quantitative analysis showed that the mean protein levels in urine before and after the addition of Evr were 87.84 ± 141.60 mg/dL and 64.31 ± 117.44 mg/dL, respectively. Post-Evr administration resulted in a highly significant decrease (27%, P = 0.00) in the mean levels (Table 3). Among the patients without proteinuria before treatment (16, 44.4%), none developed proteinuria after treatment. Of the 36 patients, 7 (19%) who had proteinuria before treatment showed no proteinuria after treatment. However, 12 patients (33.3%) continued to show proteinuria even after treatment. No reports were obtained for one patient (Table 4).

Table 3 Quantitative analysis: Proteinuria levels before and after initiation of everolimus.

Mean urinary protein (mg/dL)		SD
Before everolimus	87.84	141.60
After everolimus	64.31	117.44

P < 0.05.

Table 4 Qualitative analysis: Proteinuria levels before and after initiation of everolimus, n (%).

Parameters	Number of patients			Total patients	P value
	Before	After
Protein level not found	16 (44.4)		0 (0)	16 (44.40)	0.005a
Protein level not reported	1 (2.7)		1 (2.7)	1 (2.7)
Protein level detected	7 (19.4)		12 (33.3)	19 (52.7)
Total	24 (66.6)		13 (36.1)	36 (100)

^aP < 0.05.

Improvements in cholesterol levels were observed in 50% (n = 18) of the patients. The percentage of cholesterol level decline among patients varied from 3% to 22%. A few patients (n = 4, 11%) showed an increase of less than 10% in lipid levels, and seven (19%) showed an increase of no more than 20% after switching to Evr. This shows that the treatment maintained stable lipid levels. Of the 36 patients, only seven showed a remarkable increase in lipid levels. However, the results could not be included in the statistical analysis due to intersubject variability.

Immunosuppression

In the present study, trough levels were measured at three time points: 1 week, 6 months, and 1 year after drug administration. The trough concentrations of both drugs were maintained at the target level of 5-7 ng/mL. None of the patients showed graft rejection. There was no significant difference in the trough levels of Evr at 6 months and 1 year after 1 week. For Tac, there was a significant difference in trough levels between 1 week and after 1 year (P = 0.002) (Table 5).

Table 5 Trough levels of everolimus and tacrolimus at different time intervals.

Time	Everolimus trough levels, mean value (ng/dL)	P value	Tacrolimus trough levels, mean value (ng/dL)	P value
At week 1	3.86	0.726	4.82	0.399
Post 6 months	3.82	0.078	4.66	0.002a
Post 1 year	3.65	0.071	3.92	0.000a

^aP < 0.05.

Drug-related side effects

Side effects encountered during the MMF protocol included vomiting, urinary infection, diarrhea, and anemia. Of the total study population (n = 36), 26 patients reported vomiting, 11 suffered from diarrhea, seven developed urinary tract infection, and two presented with anemia (Figure 1). All side effects disappeared in all patients when the regimen was switched from MMF to the combination protocol of low-dose Tac + low-dose Evr.

Figure 1 Incidence of mycophenolate mofetil associated side effects. UTI: Urinary tract infections.

Safety and tolerability

No adverse events were observed in the study population during the study period. All patients achieved full recovery and survived.

DISCUSSION

Evr is an mTORi that exerts its potent immunosuppressive action by inhibiting mTOR activity, leading to the inhibition of B- and T-cell proliferation[19,20]. Evr offers an opportunity to reduce Tac doses and change the MMF regimen in kidney transplant patients, as it does not show obvious nephrotoxicity. Antiproliferative agents such as MMF and azathioprine are traditionally used in immunosuppressive regimens to prevent acute rejection and long-term deterioration of graft function[13]. However, several side effects require close monitoring. MMF is known for its GI-related side effects such as nausea, vomiting, and diarrhea. In addition to GI disturbances, it causes hematological abnormalities[19]. In the current study, all patients experienced MMF-associated side effects (vomiting, diarrhea, anemia, and urinary tract infection), and the treatment regimen was switched to Evr. After switching to Evr, none of the patients presented with similar symptoms.

Compared with MMF, Evr exhibits antiproliferative, antineoplastic, antiviral, and antiatherosclerotic properties[20]. It is particularly useful for preventing new-onset diabetes mellitus caused by other immunosuppressive agents. Corticosteroids, CNIs, and sirolimus are diabetogenic agents that cause frequent complications of post-transplant diabetes mellitus. However, these drugs are not discontinued because the IS regimen is tailored to preserve optimal kidney function rather than control diabetes[3]. By implementing CNI minimization, Evr use could potentially lower the risk of CNI-related nephrotoxicity and new-onset diabetes mellitus[21].

Evr has the potential to improve long-term outcomes but causes considerable toxicity, including proteinuria, dyslipidemia, edema, and delayed wound healing[22]. Generally, proteinuria seems to be associated with regimen conversion in maintenance patients or in those with preexisting proteinuria, rather than with de novo Evr use. Switching to Evr is not advisable for patients with proteinuria > 800 mg/day[23]. Similar criteria were used in the present study: Patients with pre-existing proteinuria > 800 mg/day were not administered Evr. However, none of the patients in this study had proteinuria > 800 mg/day. Hyperlipidemia is another side effect of Evr, and it can be managed with statins to a large extent without treatment discontinuation.

Evr also exerts non-immunosuppressive effects by facilitating mTOR inhibition. It can lead to regression of cardiac hypertrophy, inhibit atherosclerotic plaque development and rupture, and attenuate arterial stiffness[14]. In addition, the pleiotropic effects of the drug reduce the rate of post-transplant malignancy and the incidence of viral infections, thereby reducing factors that contribute to late mortality post-kidney transplant[23-26]. However, a longer follow-up of our patients is needed to determine the incidence of subsequent malignancies and late mortality.

Evr has been used in combination with other CNIs. Langer et al[27] reported the successful use of Evr with Tac minimization (low dose) in post-kidney transplant patients, who not only showed good renal function but also low graft loss rates with an acceptable safety profile. Our results are consistent with previous studies that have shown a significant reduction in serum creatinine levels after switching from MMF to Evr, indicating good renal function. Serum creatinine levels were measured before and 1 month after switching to Evr. We found a statistically significant decrease in the mean serum creatinine levels after switching to Evr (P = 0.20). However, the average serum creatinine levels evaluated at 1-year post-treatment (1.41 ± 0.44 mg/dL) were lower than the pre-Evr levels (1.45 ± 0.46 mg/dL) but were not statistically significant (P = 0.398). This indicates the long-term benefits of the combination regimen at low doses. Stabilization of creatinine levels in the treated patients indicates improved quality of life. However, identifying the most suitable Evr dose is important to achieve therapeutic blood concentrations after KT in patients receiving Tac[10,18]. Similar results were observed in our patients at 1 week, 6 months, and 1 year, revealing that we managed to maintain the recommended trough levels without significant manifestations of Evr-associated side effects.

This retrospective study assessed the efficacy of low-dose Evr and low-dose Tac in post-kidney transplant patients. Tac and Evr were administered at a dose of 0.25 mg BD. This corresponded to an eightfold lower dose than the standard drug dose. The trough concentration was defined as 5-7 mg/mL and maintained throughout the experiment. None of the patients presented with high-dose Tac- or MMF-associated side effects, probably because of the maintenance of low drug levels in the blood.

Proteinuria is a common finding in post-kidney transplant patients and is considered a clinical predictor of graft survival and mortality[28]. Although there are several causes of proteinuria following transplantation, one contributing factor is the use of immunosuppressive agents, such as mTOR inhibitors. The potential mechanisms of mTORi-associated proteinuria include reduced vascular endothelial growth factor synthesis and interference with key podocyte proteins, leading to podocyte contraction and increased permeability of the glomerular basement membrane[28,29]. In this study, urine protein levels were assessed before and after Evr administration. As stated earlier, proteinuria was defined as > 800 mg/day. After Evr administration, the protein levels were detected in only 12 patients, with the highest recorded dose being 337 mg. According to the statistical analysis, the mean proteinuria levels were significantly (P = 0.00) reduced after switching to Evr.

Proteinuria and lipid profiles were considered as indicators of graft survival in our study. No significant improvement in cholesterol levels was observed. Only half of the patients exhibited decreased cholesterol levels. Those who reported an increase also showed variable results: 11% of patients showed an increase of less than 10%, and 19% showed an increase of less than 20% in lipid levels. Our study results indicate that patients benefited from the treatment switch in terms of their lipid profiles. All patients survived, and none showed graft rejection. Furthermore, none of the patients discontinued Evr therapy, as increased cholesterol levels were successfully managed using statins[30]. However, this was a limitation of our study. Due to the high inter-subject variability (3%-22%), we were unable to perform a statistical analysis.

CONCLUSION

Our study results are consistent with previous findings, suggesting that Evr, combined with early and substantial Tac minimization and low-dose steroids, achieves good renal function and an acceptable safety profile. However, trough levels of the drugs should be monitored and maintained to reduce the possible side effects of the combination regimen used.

ACKNOWLEDGEMENTS

We acknowledge the help of Medica Superspecialty Hospital and its staff in order to carry out this study.