Advances in Immunosuppression
Immunosuppression in heart transplantation has been managed with a roughly unchanged milieu of therapy over the last decade. In order to continue the advancement of long-term patient outcomes, new therapeutic options should be explored that have enhanced efficacy or reduced toxicity over current agents as well as better ways to monitor current therapeutic options. Additionally, there has been an increased interest in modulating antibody production as a result of increased insight and experience in treating antibody-mediated rejection (AMR). This has also afforded the opportunity to explore novel therapy in patients with either AMR or elevated panel reactive antibodies prior to transplant. In this chapter, advances in maintenance therapy will be discussed including delayed-release tacrolimus, elevating the role of mammalian target of rapamycin (mTOR) inhibitors, and novel targets. Discussion of a wide variety of agents in development is also included, with a focus on effects on antibody production.
KeywordsDesensitization mTOR inhibitors Proteasome JAK inhibitor Complement
Although calcineurin inhibitors (CNI), antimetabolites, and mammalian target of rapamycin (mTOR) inhibitors significantly improved allograft and patient survival, several unmet needs still exist in the field of immunosuppression. While current therapy is efficacious at promoting excellent short-term outcomes, further research is needed to evaluate new strategies to promote enhanced long-term survival. Additionally, the current armamentarium of immunosuppressive agents carries well-known adverse effects that can further impede long-term survival, including metabolic effects (hypertension, hyperlipidemia, etc.), malignancy, and end-organ damage. According to ISHLT registry data, long-term cardiac allograft survival has a median of roughly 12 years (Lund et al. 2017). In patients with >10 years of survival, the overall frequency of malignancy is almost doubled for patients with 5 years of survival (27.7% vs. 15.9%, respectively). The frequency of cardiac allograft vasculopathy (CAV) also remains a troublesome long-term complication. Newer therapies are needed that decrease these risks and mitigate antibody-mediated allograft injury (Stegall et al. 2016). One of the major barriers to development of new therapies is the lack of novel biomarkers for use as surrogate endpoints. Most studies focus on short-term allograft and patient survival, incidence of biopsy-proven acute rejection, and safety endpoints. Although these endpoints have proven useful, they do not provide a complete picture especially since short-term outcomes have improved significantly. Moving forward, well-designed, large clinical studies would be helpful in determining optimum regimens with good long-term follow-up.
This chapter will focus on updates to current therapies, novel agents under development, potential targets for improved monitoring and decision-making, and the overall direction of transplant pharmacotherapy. It is important to note that majority of the data reviewed will be in non-cardiac allograft recipients and application to cardiac transplantation is, at best, extrapolated.
Current Maintenance Therapy
Summary of current therapies
Mechanism of action
Therapeutic drug monitoring
Calcineurin inhibitors (CNIs)
Binds to cyclophilin; complex inhibits calcineurin, IL-2-driven T-cell activation
C2 levels or C0 levels
Toxicities: nephrotoxicity, hypertension, gingival hyperplasia, hirsutism, lower incidence of neurotoxicity
Role: second-line CNI, intolerance to tacrolimus
Binds to FKBP12; complex inhibits calcineurin, IL-2-mediated T-cell activation
C0 levels, trough goal varies depending on risk of rejection, organ, time from transplant, and adjunctive therapy utilized (i.e., antimetabolites vs. PSIs)
Toxicities: similar to cyclosporine except more neurotoxicity, new-onset diabetes mellitus, alopecia, less hypertension
Updates: two novel once-daily, extended-release formulations; LCP-tacrolimus and tacrolimus ER. LCP-tacrolimus utilizes MeltDose® technology that improves bioavailability and peak-to-trough ratio
Role: first-line maintenance agent in combination with antimetabolites or proliferation signal inhibitors; potential role for LCP-tacrolimus in neurotoxicity
Proliferation signal inhibitor
Binds to FKBP12; complex inhibits mammalian target of rapamycin and IL-2-mediated T-cell activation
C0 levels, goal varies depending on monotherapy, combination with antimetabolites, or combination with CNIs
Toxicities: hyperlipidemia, thrombocytopenia, hepatic artery thrombosis, delayed wound healing, mouth ulcers, pneumonitis
Role: cardiac allograft vasculopathy, malignancy, and CMV viremia
Proliferation signal inhibitor
Derivative of sirolimus with similar mechanism of action
Binds to B7 (CD80 and CD86) receptors on antigen-presenting cells and prevents binding to CD28 on T cell which inhibits costimulation
Toxicities: high rate of rejection, PTLD (in EBV-seronegative patients), peripheral edema, hypertension, hyperkalemia, hypokalemia
Updates: renal function remained stable at 7 years and similar graft loss in comparison to cyclosporine
Role: used as backbone in patients intolerant to CNI, combination with low-dose CNI for renal sparing
One of the more notable developments in maintenance immunosuppression is the introduction of extended-release once-daily tacrolimus formulations. Currently, two extended-release formations exist – tacrolimus ER (Astagraf XL, Advagraf XL, Astellas) and LCP-tacrolimus (Envarsus XR, Veloxis). Although both formulations are dosed once daily, important differences in their formulations impact their pharmacokinetic profiles. Tacrolimus ER was developed by the addition of ethyl cellulose which slows down the diffusion rate of tacrolimus, thereby providing the prolonged release and half-life (Astragraf 2015). In contrast, LCP-tacrolimus uses a MeltDose® technology which reduces tacrolimus particle size and decreases surface area of the drug particles, thereby translating to improved solubility and bioavailability. This improvement is notable as the recommended conversion to LCP-tacrolimus is 80% of total daily dose of the immediate-release formulation (Envarsus 2017). Additionally, LCP-tacrolimus is associated with less fluctuation between peak exposure and trough concentrations, and both formulations have demonstrated noninferiority when compared with immediate-release tacrolimus. Applications for extended-release formation theoretically range from improved compliance to potential reduction in neurotoxicity. In a recent phase IIIb trial, the effect of switching stable renal transplant recipients from immediate-release to LCP-tacrolimus on tremor was studied. Thirty-eight patients were converted to LCP-tacrolimus and improvement in tremor was studied using the Fahn-Tolosa-Marin (FTM) scale, an accelerometry device, and the quality of life in essential tremor (QUEST). 78% of patients reported improvement following the switch and statistically significant reduction in FTM score as well as the accelerometer were noted (Langone 2015).
Proliferation Signal Inhibitors (mTOR Inhibitors)
The two mammalian targets of rapamycin inhibitors, sirolimus and everolimus, were introduced in 1999 and 2009, respectively. Following the advent, it was thought that these agents would replace calcineurin inhibitors and mitigate the metabolic adverse events as well as the nephrotoxicity. Unfortunately, studies have shown inferior outcomes with sirolimus as evidenced by overall higher acute rejection rates in CNI withdrawal and sparing regimens substituted with sirolimus in comparison with CNI standard of therapy (ORION, ELITE-SYMPHONY). The renal-sparing effect of sirolimus was evaluated in a randomized controlled trial in cardiac allograft recipients. Patients 1–8 years posttransplantation with mild to moderate renal impairment were either maintained on CNI-containing regimens or converted to sirolimus-based regimens. Although the mean change in creatinine clearance was significantly higher in the sirolimus arm, acute rejection associated with hemodynamic compromise and higher discontinuation rate attributable to adverse effects were also common in the sirolimus arm (Zuckermann et al. 2012). Post hoc analysis of the same cohort was conducted to determine predictors of renal function response and risk factors for acute rejection. The findings showed mycophenolate doses less than 1000 mg daily and preexisting diabetes were predictors for acute rejection and decreased renal function. It was therefore concluded that sirolimus-based therapy is an option for improving renal function in patients without preexisting diabetes and on optimal mycophenolate doses (Zuckermann et al. 2014). Additionally, delayed wound healing, proteinuria, and hyperlipidemia were more frequent in the sirolimus group. Interestingly, mTOR inhibitors are associated with lower CMV infection rates, but the utility of this property warrants further investigation (Andrassy et al. 2012). Antiproliferatives have also been shown to reduce the incidence of cardiac allograft vasculopathy (Kaczmarek et al. 2013; Mancini et al. 2003). In an open-label, prospective, randomized study of patients with documented cardiac allograft vasculopathy (CAV), a subset of patients was allotted to receive sirolimus in comparison to standard of care. It was concluded that the use of sirolimus slowed the progression of CAV and the mechanism was unlikely to be related to B-cell suppression due to lack of differences in antibody production between both groups (Mancini et al. 2003). Another large randomized, double-blind trial (n = 634) compared everolimus in combination with cyclosporine to azathioprine. Everolimus was more efficacious than azathioprine in reducing incidence of CAV (Eisen et al. 2003). Furthermore, given that increased incidence of posttransplant malignancy particularly skin cancers and the fact that the mTOR pathway has been implicated in progression of malignancies, proliferation signal inhibitors may play a role in risk reduction (Karia et al. 2016).
Belatacept, which was also designed with hopes of providing similar efficacy to calcineurin inhibitors without the associated nephrotoxicity, is a selective costimulation blocker exerts its effects by inhibiting signal 2 of T-cell activation. Since its approval in 2011, belatacept has been received with mixed reservations and is yet to find its niche due to the high risk of rejection and posttransplant lymphoproliferative disease (PTLD).
As hypothesized, the BENEFIT study, which compared belatacept to cyclosporine in renal transplant recipients, demonstrated superior renal function in the belatacept group but higher early rejection rates. Patient and graft survival at 12 months were however similar between both groups. Interestingly, belatacept groups demonstrated lower donor-specific antibody (DSA) formation rate (Vincenti et al. 2010). The BENEFIT-EXT study was restricted to recipients from extended criteria donor or cold ischemia times over 24 h (Durrbach et al. 2010). This study found 36-month rate of acute rejection, graft loss, and treatment failure was similar across all groups. Furthermore, 7-year results and post hoc analysis of the BENEFIT-EXT study showed similar time to death or graft loss between the belatacept and cyclosporine group. Estimated mean GFR increased for the belatacept group but declined in the cyclosporine group (Florman et al. 2017).
The role for belatacept in the future will be determined by identifying subpopulations that may benefit from utilizing this medication. To this end, several case studies and case series utilizing modified dosing regimens presented findings worth mentioning. A recent retrospective study reported the outcomes of conversion from tacrolimus to belatacept in high immunologic renal allograft recipient. In contrast to the BENEFIT studies, the authors included six high-risk patients defined as cPRA >80%, retransplantation, positive crossmatch at time of transplantation, and history of antibody-mediated rejection less than 3 months before the switch. Renal function improved with peak eGFR pre and post switch reported as 23.8 ± 12.9 ml/min/1.73 m2 and 42 ± 12.5 ml/min/1.73 m2, respectively. Additionally, two recipients on HD due to prolonged delayed graft function had renal recovery. Finally, overall biopsy findings improved post conversion to belatacept (Gupta et al. 2015). In the largest single-center, retrospective data to date, Adams and colleagues compared a modified belatacept regimen with a matched control group on standard tacrolimus-based maintenance regimen in kidney allograft recipients. Given the increased risk of early rejection seen in the BENEFIT studies, the group developed a modified belatacept-based regimen in combination with transient calcineurin inhibitor. Three tacrolimus groups with different trough goals and weaning duration were utilized: bela/tac short with trough goal of 8–12 ng/ml weaned off after 3 months; bela/tac extended A with trough goals 8–12 ng/ml in 3 months, 5–8 ng/ml until 6-month post-op, and 3–5 ng/ml until 9-month mark at which point tacrolimus weaning began; and lastly bela/tac extended B with trough goals 8–12 ng/ml in the first month, 5–8 ng/ml until the 6-month mark, and 3–5 ng/ml until the 9-month mark where tacrolimus weaning began. In addition to belatacept, all patients received mycophenolate mofetil 1000 mg BID and maintenance prednisone. Results showed comparable overall short-term patient and graft survival as well as improved renal function with belatacept similar to previous studies. Higher rejection rates were seen in the group modeled after the BENEFIT regimen in comparison to the historical tacrolimus cohort at 12 months (50.5% vs. 20.5%). Bela/tac short group experienced higher rejection rates versus historical tacrolimus cohort at 12 months (33/3% vs. 20.5%) but better in comparison to the low-intensity group modeled after the BENEFIT regimen. Finally, rates of acute rejection in the extended tacrolimus-belatacept regimen were similar to those seen in the historical belatacept regimen (16% vs. 20.5%). It was thereby concluded that the addition of a transient tacrolimus course to belatacept produces comparable rates of rejects and confers renal protection benefits (Adams et al. 2017).
Reported use of belatacept in cardiac transplantation is limited to one case study thus far. Briefly, a 26-year-old female heart transplant recipient secondary to postpartum cardiomyopathy was found to have erratic tacrolimus levels with raised concerns for noncompliance. The patient was noted to have several episodes of mild to moderate rejection and eventually severe grade 3R rejection with decreased ejection fraction. Following several treatments for rejection including photopheresis, high-dose methylprednisolone, addition of sirolimus, plasmapheresis, and alemtuzumab, belatacept was initiated in addition to her maintenance regimen of tacrolimus, sirolimus, mycophenolate mofetil, and prednisone. While on belatacept, both tacrolimus and sirolimus levels fluctuated and were often undetectable, biopsies remained grade 0, and echocardiography revealed normal ejection fraction. The patient expired 7 months after belatacept was initiated, but an autopsy was not performed (Enderby et al. 2014). Further evaluation of its effect is warranted before discussing broader use in heart transplant recipients.
Monitoring Advancements with Current Pharmacologic Options
Years ago, pharmacogenetics was predicted to revolutionize pharmacotherapy. Individualized medicine based on genetic profiling and anticipated effect of single nuclear polymorphisms (SNPs) on drug metabolism however has yet to become mainstay for most therapies. The initial limiting factors were cost and time required to genotype, but these tests are now affordable and readily available. Given that calcineurin inhibitors and mTOR inhibitors are metabolized by CYP enzymes, the most relevant polymorphism in the field of transplantation is in the CYP3A subfamily. Cyclosporine and tacrolimus are both substrates of CYP3A4, CYP3A5, and P-glycoprotein (ABCB1) efflux pump. Interpatient variability occurs due to difference in expression as well as mutations in the genes encoding these enzymes (Staatz et al. 2010).
Interestingly, it has been reported that race plays a role in these polymorphisms which results in varied dose requirements between white and black allograft recipients. The loss-of-function allele CYP3A5∗3, which is the major allele in whites, has been identified as a major determinant of variation in tacrolimus metabolism due to reduction in activity which results in higher drug concentrations. The loss of activity results from alternate splicing of the third intron of the CYP3A5 gene which leads to an out-of-frame mRNA variant that codes for a nonfunctional protein. Conversely, persons with CYP3A5∗1 allele metabolize tacrolimus rapidly resulting in overall lower whole blood concentrations and higher dose requirements. Additionally, a recent genomewide association study of tacrolimus trough in black kidney allograft recipients identified two novel CYP3A5 variants that were associated with tacrolimus troughs. The two loss-of-function alleles CYP3A5∗6 and CYP3A5∗7 were found to play significant roles in tacrolimus metabolism (Oetting et al. 2016).
The role of CYP3A5 genotype on dose requirement of tacrolimus and everolimus was evaluated in white cardiac allograft recipients by Kniepeiss and colleagues. In this study, 15 patients received tacrolimus-based regimens, whereas 30 patients received everolimus-based regimens. Thirteen subjects in the tacrolimus group were CYP3A5 non-expressers, i.e., homozygous for ∗3 allele, whereas two were heterozygous expressers. When compared, average tacrolimus dose requirement was significantly higher in the expressers. In the everolimus group, 27 patients were homozygous for ∗3 allele, and 3 were heterozygous. Interestingly, there was no significant difference in dose or levels between expressers and non-expressers. It was thereby concluded that CYP3A5 polymorphism affects tacrolimus dose requirement but the same influence is not seen with everolimus (Kniepeiss et al. 2011). Most recently, the impact of CYP3A5 genotype on extended-release formulation of tacrolimus (LCP) was studied in 50 black renal allograft recipients. Eighty percent were CYP3A5 expressers, and there were no significant differences in AUC between expressers with immediate-release and LCP-tacrolimus. Interestingly, Cmax was 33% higher with the immediate-release formulation in CYP3A5 expressers in comparison to 11% higher with LCP-tacrolimus. It was concluded that this difference in peak could theoretically reduce peak-related toxicities (Trofe-Clark et al. 2017).
In conclusion, determination of CYP3A5 polymorphism in combination with therapeutic drug monitoring potentially reduces time to therapeutic levels (MacPhee et al. 2004). The clinical benefit in terms of allograft function and rejection of this strategy however is yet to be established.
Immune Cell Function Monitoring
Immune cell function monitoring, FDA approved in 2002, takes a unique approach to monitoring patients on immunosuppressive regimens. This assay, commercially known as ImmuKnow, utilizes lymphocyte activation to ascertain a net state of immune function. Lymphocytes are stimulated with phytohemagglutinin (PHA), thus resulting in an increase in cellular energy supply and utilization. This assay then determines the amount of intracellular adenosine triphosphate (ATP) in lymphocytes upon activation. Given that CD4 cells are the target for major immunosuppressive drugs like cyclosporine and tacrolimus, this cell line was chosen by investigators to undergo ATP measurements (Kowalski et al. 2003).
Kowalski and colleagues evaluated this monitoring modality in 127 transplant patients (compared with healthy individuals) to attempt to determine if breakpoints exist where patients may be over- or under immunosuppressed. Patients were determined to have a “low immune response” if ATP levels were ≤225 ng/ml, whereas “strong immune response” was defined as ATP levels ≥525 ng/ml; 94% of healthy volunteers had values ≥225 ng/ml versus 92% of transplant patients had values ≤525 ng/ml. When patients were analyzed depending on the immunosuppressant used, there was no difference in ATP levels between patients treated with cyclosporine versus tacrolimus. The authors did note that drug levels did not correlate with ATP levels, citing the importance of assessing overall immune function not just readily available therapeutic drug monitoring (Kowalski et al. 2003).
Kobashigawa and colleagues evaluated the clinical utility of this assay in heart transplantation in a single-center study (Kobashigawa et al. 2010). Patients had ATP assays ranging from 1 month to 10 years after transplant and were treated with tacrolimus, mycophenolate, and corticosteroid without induction therapy. The authors’ goal was to determine if breakpoints existed to identify patients at risk for either infection or rejection based upon their ATP results. Patients that developed infections had a statically lower ATP level than patients at steady state without infections (187 ± 126 ng ATP/ml vs. 280 ± 126 ng ATP/ml, p < 0.001). There was no difference in mean ATP levels in steady-state patients versus those with rejection, but this could have been due to the low frequency of rejection overall in this study population. These results with respect to infection were consistent with what has been reported in other trials (Kowalski 2006; Thai et al. 2006).
Immune function monitoring has had more recent data published examining its utility in the management of transplant patients. Ben Gal and colleagues evaluated immune monitoring (IM) in 34 heart transplant recipients managed with everolimus-based immunosuppression (Ben Gal et al. 2014). Patients had a wide variance in follow-up periods, with samples being drawn at 1 week after transplant. ATP levels obtained during infection episodes were significantly lower when compared to patients who were not infected (188 ± 122 ng/ml vs. 338 ± 137 ng/ml; p < 0.05). Again, there was no difference in values between patients with and without rejection.
Ravaioli and colleagues evaluated immune function monitoring in a novel way by using it to aid in adjusting tacrolimus levels in liver transplant recipients (Ravaioli et al. 2015). Liver transplant recipients from July 2008 to March 2013 were randomized to standard care (n = 102) described as tacrolimus-based immunosuppression with oral steroids. Goal tacrolimus levels for standard care were 8–12 ng/ml for the first 4 months and 6–10 ng/ml thereafter. Patients who had therapy guided by immune function monitoring had goal tacrolimus levels reduced by 25% when ATP levels <130 ng/ml and increased by 25% when ATP levels were >450 ng/ml (n = 100). Patient survival was higher at 12 months in the intervention group (89% vs. 80%, p < 0.05). Additionally, patients managed with immune function monitoring had less infection episodes (42% vs. 56%, p < 0.05). Rejection episodes were not different between groups. The authors concluded that goal-level adjustments with the immune function assay were beneficial in this patient population by providing additional data on the net state of immunosuppression.
Immune function monitoring can be of assistance in patients at risk for infectious complications, especially in a low-rejection risk patient population. Results from clinical trials have not shown the same benefit with respect to rejection reduction. This is mostly due to low rejection numbers in these reports. Immune function monitoring does provide additional information with respect to the net state of immunosuppression that cannot be ascertained by drug-level monitoring alone. Use of these advanced monitoring techniques, as well as pharmacogenomics analysis, can help transplant centers manage patients in a more precise fashion. Pharmacogenomics can assist in determining upfront dosing strategies by identifying metabolic differences among patients. When immune function monitoring is utilized to guide traditional therapeutic drug monitoring strategies, it appears that we can reduce infectious complications of transplantation which is a major obstacle to avoid in providing successful outcomes in this patient population.
Investigational Agents for Maintenance Immunosuppression
Emerging immunosuppressants currently in development phase
Mechanism of action
Place in therapy
Costimulation (CD40–CD154) inhibitor
Inhibits humoral and cellular immune responses by blocking the CD40/CD154 interaction between antigen-presenting cells and T cells
Similar efficacy outcomes at 6 months when combined with low-dose tacrolimus but inferior when combined with MMF
Janus kinase inhibitor
Inhibits JAK/signal and transducers and activators of transcription which prevents T-cell proliferation
Similar BPAR at 6 months in comparison to cyclosporine but higher incidence of anemia, PTLD, and viral infections associated with excessive immunosuppression in the tofacitinib group
Reversible inhibition of the 26S proteasome resulting in plasma cell death
Peripheral neuropathy (dose dependent), thrombocytopenia, neutropenia, neuralgia, gastrointestinal toxicity, CYP3A4-mediated drug interactions
Irreversible inhibition of the 26S proteasome resulting in plasma cell death
Acute renal insufficiency, venous thrombosis, hypertension, thrombocytopenia, no CYP-mediated drug interactions
Binds C5 protein of the complement cascade, thereby preventing formation of the membrane attack complex
FDA approved for atypical hemolytic uremic syndrome and paroxysmal nocturnal hemoglobinuria. Due to life-threatening meningococcal infections, meningococcal vaccines required at least 2 weeks prior to the first dose of eculizumab
Toxicities include hypertension, tachycardia, rash, headache, diarrhea, nausea and vomiting, and upper respiratory infection
C1 esterase inhibitor
Inhibits complement proteins C1r and C1s through which the classical complement pathway is interrupted
Toxicities include headache, abdominal pain, and oropharyngeal pain
B-lymphocyte stimulator (BLyS) protein inhibitor
Prevents BLyS from binding to receptors on B cells which reduces B-cell-mediated immunity
FDA approved for systemic lupus erythematous. Toxicities include nausea, diarrhea, infusion-related reaction, and hypersensitivity
IL-6 inhibition inhibits B-cell progression to plasma cell
FDA-approved rheumatoid arthritis and idiopathic juvenile arthritis
In a phase II trial, TCZ used in combination with IVIg and rituximab allowed the transplantation of 5 highly sensitized patients
IgG endopeptidase (IdeS)
Bacterial IgG proteinase
Bacterial IgG isolated from Streptococcus pyogenes proteinase that cleaves all 4 human IgG subtypes
In a phase II trial, 24 of 25 highly sensitized patients successfully underwent renal transplantation from HLA-incompatible donors
In a phase Ib study, the efficacy and safety of ASKP1240 were evaluated in 138 renal transplant recipients stratified to either a calcineurin-free regimen containing solely of the study drug and MMF (n = 46) versus study drug in combination with reduced-intensity tacrolimus (n = 44) versus standard of care which comprised of tacrolimus and MMF (n = 48). With respect to safety, none of the subjects experienced thromboembolic events or PTLD; however, three patients developed malignancies, and higher rates of viral infections were reported in the study group. It was concluded that ASKP1240 in combination with reduced-intensity tacrolimus attained similar efficacy at 6 months, while similar efficacy was not achieved in the ASKP1240 + MMF group (Harland et al. 2015). Given the limited efficacy as backbone immunosuppression and increased incidence of viral infections, the utility, as well as the future, of ASKP1240 remains unclear. CFZ533 is another antiCD40 fully human monoclonal antibody that is currently being evaluated for safety and efficacy (NCT02217410).
JAK Inhibition with Tofacitinib
As an overview, JAKs are tyrosine kinases that facilitate signal transduction and activators of transcription (STAT) phosphorylation, dimerization, and nuclear transport which results in transcription and gene expression. There are four mammalian JAK subtypes, namely, JAK1, JAK2, JAK3, and tyrosine kinase 2 (Wojciechowski 2013). Unlike other subtypes, JAK3 is found primarily on hematopoietic cells, and the importance in immunosuppression has been demonstrated through murine models. Mice that lacked common gamma chain (cy) of cytokines or JAK3 were found to develop severe combined immunodeficiency syndrome (Wojciechowski 2013). Additionally, tofacitinib is 20- to 100-fold less potent for JAK2 and JAK1 which could translate to less hematologic adverse effects such as anemia, thrombocytopenia, and leukopenia (Wojciechowski 2013).
Tofacitinib, formerly known as CP-690,550, is an orally active Janus kinase (JAK) inhibitor that is FDA-indicated for treatment of moderate to severe rheumatoid arthritis (Xeljanz 2015). In the tri-signal model of T-cell activation, induction of signals 1 and 2 lead to expression of cytokines such as IL-2 and IL-15. These cytokines subsequently activate the mammalian target of rapamycin via phosphatidylinositol 3-kinase and the JAK/STAT signal transduction pathway. Activation of the final signal transduction, in combination with de novo nucleotide synthesis, results in lymphocyte proliferation constituted by signal 3. By inhibiting the JAK/signal transducers and activators of transcription (STAT) pathway, signal 3 of T-cell activation is terminated. Thus, it has been theorized that tofacitinib offers an alternative to CNI-based protocols.
To date, two clinical trials have examined the safety and efficacy of tofacitinib when used for rejection prophylaxis. The first trial which was a phase IIa pilot study compared two doses of tofacitinib 15 mg (n = 20) and 30 mg twice daily (n = 20) versus tacrolimus (n = 21) in renal transplant recipients (Busque et al. 2009). In brief, patients received IL-2 receptor antagonist induction, mycophenolate mofetil, and corticosteroids. Biopsy-proven acute rejection (BPAR) after 6 months was 5 [−8.4, 9.4], 21 [2.9, 29.7], and 4.8% for 15 mg twice daily, 30 mg twice daily, and tacrolimus groups, respectively. Furthermore, the risk of BK virus nephropathy and cytomegalovirus was higher in patients treated with tofacitinib when compared to patients treated with tacrolimus. This study concluded that 15 mg twice daily was the preferred dose as 30 mg twice daily resulted in excessive immunosuppression without benefit in allograft or patient survival. In a phase IIb study, the second trial compared the efficacy and safety of tofacitinib at two different dosing strategies to cyclosporine in renal transplant recipients (Vincenti et al. 2012). Similar to the previous trial, all patients received basiliximab induction, mycophenolic acid, and corticosteroids. Similar BPAR at 6 months was observed for all groups, but anemia, neutropenia, and posttransplant lymphoproliferative disorder (PTLD) occurred more frequently in the tofacitinib groups. This study concluded that when combined with mycophenolic acid, tofacitinib is effective in preventing allograft rejection, demonstrated a beneficial effect on renal function, but exposes patients to higher risk of excessive immunosuppression as evidenced by incidences of serious infections, opportunistic infection, and PTLD (Vincenti et al. 2012). Finally, a post hoc analysis of the second study aimed to determine if a patient subgroup with an acceptable risk-benefit profile could be identified based on median exposure. Higher 2-hour post-dose levels (C2) correlated with the incidence of serious infection, but leukopenia and neutropenia remained significant in both the above-median exposure group and the below-median group. The findings from this study raise the potential role of therapeutic drug monitoring in minimizing toxicities while retaining efficacy (Vincenti et al. 2015; Moore et al. 2017).
Desensitization and Antibody-Mediated Rejection
Allosensitization can occur due to pregnancy, blood transfusion, and mechanical circulatory support devices. Rates of transplantation for sensitized patients remain low due to known increased risk for antibody-mediated rejection (AMR) and poor graft and patient outcomes. Approximately 20% of patients that receive heart transplants have an elevated panel reactive antibody (PRA >10%) at the time of transplant (Lund et al. 2017). Traditional efforts to reduce sensitization to HLA antigens, also known as desensitization, include plasmapheresis, IVIg, and rituximab. Although often successful, these therapies remove circulating antibodies but have negligible effect on plasma cells. Due to this, a significant percentage of the sensitized patients are refractory to standard of therapy; hence, novel therapies aimed at alternate mechanisms are crucial. Furthermore, antibody-mediated rejection is often difficult to treat and is treated using similar agents. Finally, it is important to note that there is a paucity of well-designed studies to guide therapy in AMR in heart transplant recipients. While consensus definitions exist for AMR diagnosis, there remains a clinical conundrum with respect to patients with circulating DSA without overt signs of graft damage or graft dysfunction. Currently, there are no FDA-approved agents for desensitization or treatment of AMR. This section will focus on emerging therapies and the future of desensitization as well as antibody-mediated rejection.
Proteasome inhibitors are FDA approved for the treatment of myeloma. There are three proteasome inhibitors currently available – bortezomib and carfilzomib which are available as intravenous formations and, most recently, ixazomib which is available as an oral formulation. To appreciate the utility of proteasome inhibitors, understanding the role of the proteasome is crucial. The 26S proteasome is a multi-catalytic enzyme expressed in the cytoplasm of eukaryotic cells. Its primary function is targeted degradation of ubiquitin-labeled misfolded proteins, cell cycle regulatory proteins, transcription factors, and inhibitory molecules (Walsh et al. 2012). Plasma cells produce antibodies, a proportion of which will be misfolded and ultimately require degradation by proteasome complex. Proteasome inhibition therefore disrupts homeostasis ultimately resulting in apoptosis. Other mechanisms through which proteasome inhibitors exert immunomodulatory effects include inhibition of class I MHC expression through reduction of endogenous peptide production, as well as inhibition of nuclear factor kappa B (NF-kB) activity which leads to subsequent reduction in cytokine production including IL-6 (Sadaka et al. 2012). Bortezomib is a first-in-class proteasome inhibitor that binds the 26S proteasome reversibly, whereas carfilzomib is a second-generation proteasome inhibitor that binds irreversibly. When compared, bortezomib has potential for neurotoxicity and drug-drug interactions through CYP3A4 and 2C19, whereas carfilzomib has lower incidence of neurotoxicity and no CYP-mediated drug interactions (Velcade PI, Kyprolis PI). Although they are not FDA approved for AMR or desensitization, these agents are being used in refractory cases in combination with standard of care. Ixazomib gained FDA approval in 2015 and like bortezomib is a reversible proteasome inhibitor (Ninlaro PI 2017), but it has yet to be used in transplantation. Like other modalities, proteasome inhibitors have been studied mainly in renal allograft with a few case series emerging in cardiac allograft recipients. In a pilot study, bortezomib was utilized in six allosensitized patients awaiting cardiac transplantation with persistently elevated anti-HLA antibodies despite conventional therapy with IVIg and rituximab. Mean calculated panel reactive antibody (cPRA) reduced from 62% to 35%, but infection was common after treatment (Patel et al. 2011). A recent case series described the use of carfilzomib in 14 lung allograft recipients with AMR. The regimen included plasma exchange and IVIg in combination with carfilzomib. Patients were deemed responders if complement-1q (C1q) was suppressed after treatment. Seventy-one percent responded to carfilzomib and had less chronic lung allograft dysfunction or progression when compared to nonresponders (Ensor et al. 2017). Moving forward, the role of proteasome inhibitors may not be limited to AMR and desensitization. Several ongoing studies are evaluating the utility in induction strategies as well as chronic rejection. Given the current body of evidence and clinical experience, proteasome inhibitors have been added to centers’ regimens as agents for desensitization and antibody-mediated rejection. Their place in therapeutic algorithms remains to be determined due to the lack of large-scale clinical trial data.
B-lymphocyte stimulation (BLyS) family of cytokines can regulate clonal selection and the B-cell lifespan (Parsons et al. 2010). Belimumab exerts its effect by preventing BlyS protein from stimulating B-cell activation and differentiation (Sethi et al. 2017). It is FDA approved for systemic lupus erythematous. Currently, induction and maintenance therapy are T lymphocyte directed with negligible effects on B lymphocyte. B lymphocytes have been implicated in chronic antibody-mediated rejection; thus, exploring depletion at the time of transplant could be beneficial in both allosensitized and non-sensitized patients (Parsons et al. 2010). A recent case report demonstrated the potential efficacy of belimumab for AMR in a kidney-pancreas recipient. Briefly, the patient developed AMR associated with high levels of HLA-DR53 antibodies which was resistant to treatment with plasmapheresis, IVIg, and rituximab. Following treatment with belimumab, HLA-DR53 mean fluorescence intensity (MFI) decreased by 30%, and serum creatinine decreased from 4.5 to 2.8 mg/dl. It was thereby concluded that belimumab could be an effective therapy for AMR (Leca and Muczynski 2013). A clinical trial aimed at examining the effect of belimumab as a desensitizing agent in kidney transplantation was terminated due to lack of efficacy (NCT01025193). Another phase 2 trial examining belimumab for rejection prophylaxis in combination with standard of therapy is currently ongoing (NCT01536379).
Eculizumab (C5 Inhibitor)
Complement activation is another interesting target in the field of transplantation, with the ultimate formation of the membrane attack complex resulting in devastating cellular injury. Eculizumab is a humanized monoclonal antibody that exerts its effect by binding to C5 protein of the complement cascade, thereby preventing formation of the membrane attack complex. It carries the FDA indication for atypical hemolytic uremic syndrome (aHUS) and paroxysmal nocturnal hemoglobinuria (PNH). Most studies in transplantation have been in renal allografts for treatment and prevention of AMR. In an open-label trial, the effect of eculizumab in addition to plasmapheresis and thymoglobulin on AMR 3 months posttransplant in 26 highly sensitized renal allograft recipients (defined as positive crossmatch against their donors) was evaluated. The study group was compared to a historical highly sensitized group (n = 51) who received similar plasmapheresis-based protocol. Incidence of AMR was significantly lower in the study group (7.7% vs. 41.2%, p = 0.0031). It was thereby concluded that eculizumab decreases the incidence of early AMR in sensitized renal allograft recipients (Stegall et al. 2011). A nonrandomized, open-label study investigating the use of eculizumab in combination with conventional therapy in highly sensitized cardiac transplant patients is currently ongoing. This study aims to determine if this strategy will prevent antibody-mediated rejection and prolong long-term cardiac allograft survival (NCT02013037). Interestingly, eculizumab may not be effective in c4d-negative AMR as evidenced by two case reports in renal transplant recipients (Burbach 2014; Tran 2016). Since c4d is a sign of complement activation, the lack of efficacy of eculizumab in c4d AMR cases suggests other mechanisms may be involved in AMR. Finally eculizumab is an expensive treatment and is not without risk. Specifically, due to increase in incidence of life-threatening and fatal meningococcal infections with eculizumab use, patients must be immunized with meningococcal vaccines at least 2 weeks prior to administering the first dose (Soliris PI). Given the economic consideration as well as the risk for infection, most centers employ eculizumab strictly on a case-by-case basis for both desensitization and treatment of antibody-mediated rejection.
C1 Esterase Inhibitor
C1 esterase inhibitor (C1-INH) is an investigation agent that inhibits complement proteins C1r and C1s. Through this inhibition, activation of the classical complement pathway is interrupted. Additionally, C1-INH inhibits mannose-binding lectin pathway through inhibition of the serine protease. In a recent phase I/II study completed with the aim of preventing antibody-mediated rejection in highly sensitized kidney transplant recipients, no antibody-mediated rejection was seen in the C1-INH group during the study duration in comparison to one AMR in the control group during the study and two afterward. It was concluded that C1-INH may be useful in preventing AMR following transplantation in highly sensitized patients (Vo et al. 2015a).
Tocilizumab (TCZ) is an IL-6 and binds both soluble and membrane-bound IL-6 receptors antagonist that carries the FDA approval for treatment of moderate to severe rheumatoid arthritis (RA) and idiopathic juvenile arthritis. IL-6 is a major cytokine involved in B-cell progression to plasma cells. Targeting this pathway leads to reduction of plasma cells and ultimately antibody production which has potential for both desensitization and antibody-mediated rejection (Jordan et al. 2015). A recent phase I/II single-center, open-label, exploratory study examined the safety and limited efficacy of the addition of TCZ to IVIg for desensitization in highly sensitized patients unresponsive to IVIg and rituximab. Ten patients were enrolled, five were transplanted, and 6-month protocol biopsies showed no antibody-mediated rejection. It was concluded that the addition of TCZ to IVIg for desensitization appears to be safe and a possible alternative in highly sensitized patients refractory to standard therapy (Vo et al. 2015b). Large, randomized controlled studies are required to confirm these findings before the utility of TCZ in transplantation can be confirmed.
IgG Endopeptidase (IdeS)
IdeS (IgG endopeptidase) is a bacterial IgG proteinase isolated from Streptococcus pyogenes. IgG endopeptidase cleaves all four subtypes of human IgG with high specificity (Björck 2016). This presents a novel avenue for desensitization as proteolytic cleavage of IgG molecules should prevent IgG-mediated antibody-dependent cytotoxicity as well as complement-mediated cytotoxicity. In 2 recent phase II trials evaluating the safety and efficacy of IdeS for desensitization, 25 highly sensitized patients received study drug prior to renal transplantation from HLA-incompatible donors. Reduction or elimination of DSAs permitted successful transplantation in 24 of 25 patients. Antibody-mediated rejection occurred in ten patients who were all responsive to treatment, and one graft loss mediated by non-HLA IgM and IgA antibodies occurred. It was concluded that IdeS reduced or eliminated DSAs at time of transplantation but it did not prevent reconstitution of the DSAs (Jordan et al. 2017).
Over the last 50 years, the field of cardiac transplantation has continually evolved from fringe medical science to an incredibly viable option for thousands of patients with advanced heart failure. For outcomes to continue on the current trajectory, development and research must exist to both optimize the way currently available agents are utilized as well as discovering improved therapeutic targets. While the majority of recent advances in transplant therapy have been for desensitization and antibody-mediated rejection, these advances borrow from other immunologic focused fields such as oncology and rheumatology. Agents such as bortezomib and carfilzomib, although still investigational, are gaining more ground as standard of therapy. Conversely, other therapies such as belimumab, c1 esterase inhibitors, IgG endopeptidase, and eculizumab are still finding their niche. Expanded understanding of mTOR inhibition, especially combined with other conventional therapies, will aid in fully harnessing the benefits of this novel class of agents. Lastly, continuing to develop improved formulations to enhance patient compliance is an important, if not overlooked, aspect to advancing cardiac transplant care. Continued focused efforts should aim to both better define the roles of our current agents and expand the number of agents available to clinicians and ultimately our patients.
- Astragraf (tacrolimus XL) (2015) [Package Insert]. Astellas Pharma US, NorthbrookGoogle Scholar
- Envarsus (tacrolimus ER) (2017) [Package Insert]. Veloxis Pharmaceuticals, EdisonGoogle Scholar
- Harland R, Klintmalm G, Yang H et al (2015) ASKP1240 in de novo kidney transplant recipients [abstract]. Am J Transplant 15(Suppl 3). http://atcmeetingabstracts.com/abstract/askp1240-in-de-novo-kidney-transplant-recipients/. Accessed 28 Oct 2017
- Kaczmarek I, Zaruba MM, Beiras-Fernandez A et al (2013) Tacrolimus with mycophenolate mofetil or sirolimus compared with calcineurin inhibitor-free immunosuppression (sirolimus/mycophenolate mofetil) after heart transplantation: 5-year results. J Heart Lung Transplant 32:277–284PubMedCrossRefGoogle Scholar
- Leca N, Muczynski K (2013) Belimumab (anti-BAFF/BLyS) effective in a case of resistant antibody mediated rejection [abstract]. Am J Transplant 13(Suppl 5). http://atcmeetingabstracts.com/abstract/belimumab-anti-baffblys-effective-in-a-case-of-resistant-antibody-mediated-rejection/. Accessed 20 Dec 2017
- Trofe-Clark J, Brennan DC, West-Thielke P et al (2017) Results of ASERTAA, a randomized prospective crossover pharmacogenetic study of immediate-release versus extended-release tacrolimus in African American kidney transplant recipients. Am J Kidney Dis 20:1–12Google Scholar
- Xeljanz (tofacitinib) (2015) [Package Insert]. Pfizer Labs, New YorkGoogle Scholar