Cyclosporine in hematological disorders: mechanisms, clinical practice and emerging advances

Abstract

Cyclosporine A (CsA) functions as a calcineurin inhibitor that perturbs T cell activation via calcineurin-nuclear factor of activated T cells (CaN-NFAT) signaling inhibitors, establishing its central role in hematological therapeutics. This review delineates the contemporary applications and mechanistic underpinnings of CsA across acquired bone marrow failure syndromes spanning severe and non-severe aplastic anemia (AA), myelodysplastic neoplasms, graft versus host disease (GVHD), and autoimmune cytopenias. We underscore individualized treatment algorithms steered by molecular biomarkers such as STAT3 mutational status, T cell receptor clonality, and telomere attrition, alongside the imperative of therapeutic drug monitoring at trough (C0) and 2-hour post-dose (C2) intervals to refine risk to benefit profiles. The manuscript further elaborates on pharmacological synergies between CsA and novel targeted agents including eltrombopag, ruxolitinib, and immune checkpoint inhibitors, while evaluating its capacity to surmount chemotherapeutic resistance and function as a bridging modality to CAR-T cell infusion. Lastly, we propose tiered management protocols for dose-limiting toxicities (nephrotoxicity and hypertension) and highlight emerging research frontiers in nanoformulation and artificial intelligence-guided therapeutic drug monitoring.

Introduction

Since its discovery in the 1970s, CsA has emerged as a cornerstone immunosuppressive agent in organ transplantation and autoimmune disease management [1]. As a calcineurin inhibitor (CNI), CsA forms a complex with intracellular cyclophilin to specifically block the CaN-NFAT signaling cascade, thereby suppressing T-cell activation and the transcription of pro-inflammatory cytokines such as interleukin-2 (IL-2) and interferon-γ (IFN-γ). This mechanism establishes its central role in treating immune-mediated hematological disorders. Unlike glucocorticoids, which exert broad genomic effects through glucocorticoid receptor-mediated transactivation and transrepression of multiple inflammatory genes, the precise T-cell regulation afforded by CsA confers unique therapeutic value in conditions including aplastic anemia (AA), myelodysplastic syndromes (MDS), graft-versus-host disease (GVHD), and autoimmune blood diseases [2]. However, the application of CsA in hematological malignancies presents a notable duality: while its immunosuppressive properties may compromise anti-tumor immune surveillance, CsA can exert therapeutic effects in specific myeloid neoplasms by modulating the hematopoietic stem cell microenvironment, reversing P-glycoprotein-mediated multidrug resistance, or stabilizing the Treg/Th17 equilibrium [3].

In recent years, deeper understanding of CsA pharmacokinetics and individual metabolic variability, coupled with the emergence of novel therapeutic agents, including eltrombopag, Immune Checkpoint Inhibitors (ICIs), JAK inhibitors, and targeted therapies, has fundamentally transformed the therapeutic landscape of CsA in hematology. Personalized therapeutic drug monitoring (TDM), precision stratification based on biomarkers (such as STAT3 mutation status, TCR repertoire oligoclonality, and telomere length), and optimized low-dose combination strategies offer new possibilities for balancing efficacy and safety. This review aims to systematically summarize the latest evidence-based data regarding CsA in acquired bone marrow failure syndromes, the management of hematopoietic stem cell transplantation (HSCT)-related complications, particularly graft-versus-host disease (GVHD) prophylaxis and treatment, autoimmune hematological diseases, and hematological malignancies. We delve into its molecular mechanisms, TDM strategies, adverse event management, and synergistic applications with novel therapies, thereby providing theoretical grounding and practical guidance for the precision and individualized use of CsA in clinical practice.

Chemical structure and mechanisms of action

Molecular structure and pharmacokinetic properties

CsA is a cyclic undecapeptide (molecular weight 1202.6 Da) composed of 11 amino acids, originally isolated from the fungus Tolypocladium inflatum [4]. Its richness in N-methylated amino acids confers lipophilicity and transmembrane capability [5]. Unlike the tetracyclic steroidal structure of glucocorticoids, CsA exerts its function by binding to intracellular cyclophilin. However, the clinical utility of CsA is substantially hampered by its erratic pharmacokinetic profile. Oral bioavailability displays considerable interindividual variability (approximately 20–50%), primarily attributable to intestinal P-glycoprotein (P-gp) efflux activity and extensive first-pass metabolism [6]. Table 1 provides a comprehensive comparison of the physicochemical and pharmacological properties of commonly employed immunosuppressive agents.

Table 1.

Comparison of Characteristics of Commonly Used Immunosuppressive Agents

Drug	Molecular Target	Mechanism	Primary Applications in Hematology	Key Toxicities	Onset of Genomic Effects	Impact on Immunotherapy	Monitoring Parameters
CsA	Cyclophilin-CaN	Inhibition of NFAT dephosphorylation	AA, MDS, GVHD	Nephrotoxicity, hypertension	Slow (2–4 weeks)	Compatible at low doses	C0 or C2 monitoring
Tacrolimus	FKBP12-CaN	Inhibition of NFAT dephosphorylation	GVHD prophylaxis	Neurotoxicity, hyperglycemia	Slow (2–4 weeks)	Compatible at low doses	C0 monitoring
Glucocorticoids	Glucocorticoid receptor	Genomic/non-genomic effects	ALL, lymphoma, ITP	Osteoporosis, metabolic disorders	Rapid (24–72 h)	Significantly impairs ICI efficacy	Clinical symptom assessment
Mycophenolate mofetil	IMPDH	Inhibition of T/B cell proliferation	Second-line aGVHD therapy	Myelosuppression, infection	Slow (4–8 weeks)	Moderate impact	Blood concentration monitoring

Core immunosuppressive mechanism

The primary immunosuppressive action of CsA begins with its intracellular binding to cyclophilin. This complex then inhibits the phosphatase activity of CaN. As CaN is essential for dephosphorylating NFAT transcription factors, its inhibition prevents NFAT from translocating to the nucleus. This blockade halts the transcription of key pro-inflammatory cytokines, including IL-2 and IFN-γ [7, 8]. In the pathological context of AA, CsA not only blocks NFAT-mediated IL-2 transcription but also rectifies Th1/Th2 and Th17/Treg cellular imbalances [9, 10]. The CsA’s impact on Treg function demonstrates concentration-dependent characteristics: at therapeutic concentrations (150–250 ng/mL), CsA selectively preserves Treg immunosuppressive function, whereas at high concentrations (> 400 ng/mL), it broadly suppresses all T-cell subsets [11]. This concentration-dependent effect on Tregs is central to a broader clinical challenge: optimizing CsA therapy. Considerable debate persists over ideal target concentrations, with emerging evidence indicating that the optimal range varies significantly based on the specific disease, concomitant medications, and treatment phase.

In AA, the classic target trough range of 150–250 ng/mL is derived from landmark immunosuppressive therapy (IST) trials [12]. Contemporary protocols have refined this approach; for instance, one recent trial specifies a target trough of ~ 150 ng/mL for patients with normal renal function, necessitating dose reduction to 25–50 mg every 12 h for those with renal impairment [13]. This pragmatic adjustment balances efficacy against the risk of nephrotoxicity. Conversely, pediatric-specific data from the IWK Health Centre recommend a higher target of 200–400 ng/mL for AA, underscoring significant age-related pharmacokinetic differences [14].

For GVHD prophylaxis following allogeneic HSCT, target concentrations are particularly contentious. While traditional recommendations suggest 200–400 ng/mL, a 2022 retrospective study of 399 myeloablative transplant recipients challenged this paradigm. Using receiver operating characteristic analysis, Kwan et al. identified 250 ng/mL as the optimal cutoff for predicting grade II–IV aGVHD; patients with mean trough concentrations < 250 ng/mL during days 15–28 post-transplant had significantly higher aGVHD rates (31.5% vs. 18.8%; OR, 1.97) [15]. Importantly, no significant association was found between CsA levels and relapse or non-relapse mortality, suggesting that maintaining higher concentrations within the therapeutic window may safely improve outcomes.

In MDS, the evidence base is even less definitive. A Phase II trial of lenalidomide plus CsA targeted a broad trough range of 100–450 ng/mL, acknowledging the uncertainty surrounding optimal immunosuppression in this heterogeneous disease [16]. The lower bound of this range reflects concern that excessive calcineurin inhibition might impair anti-tumor immune surveillance in patients with clonal disorders, while the upper bound aims to achieve meaningful suppression of pathological T-cell responses in immune-mediated cytopenias. This finding provides theoretical support for CsA application in pre-malignant conditions such as MDS, where stabilizing the Treg/Th17 balance may delay clonal evolution. Table 2 illustrates the therapeutic targets and predictive biomarkers of efficacy for CsA across various hematological disorders.

Table 2.

Therapeutic Targets and Efficacy Predictive Biomarkers of CsA cross Different Hematological Disorders

Disease Type	Pathological Mechanism	CsA Therapeutic Target	Efficacy Predictive Biomarkers	Target Trough Concentration (ng/mL)
SAA [17–20]	Th1/Th17-mediated HSPC destruction	Inhibition of CaN-NFAT, inhibitors of IFN-γ/IL-17	Telomere length > 5 kb, wild-type STAT3	150–250
NSAA [21]	Treg functional deficiency	Preservation of Tregs, suppression of effector T cells	Normal TCR Vβ repertoire	100–200
Low-risk MDS [21, 22]	Th17/Treg imbalance with STAT3 mutations	Suppression of pathological Th17 responses	STAT3 mutation+, TCR oligoclonality	150–250
SR-aGVHD [23, 24]	Multi-pathway immune activation	CaN-NFAT + JAK-STAT dual inhibition	No specific biomarkers required	200–400
Steroid-resistant ITP [25, 26]	Autoantibodies + T-cell abnormalities	Suppression of T-B cell cooperation	↓CD8 + T cell activation markers	150–250

Note: SAA = Severe Aplastic Anemia; NSAA = Non-Severe Aplastic Anemia; SR-aGVHD = Steroid-Refractory Acute Graft-versus-Host Disease; ITP = Immune Thrombocytopenia; HSPC = Hematopoietic Stem/Progenitor Cell.

Note: Target trough concentrations represent clinical practice guidelines, rather than absolute thresholds and may vary based on transplant type (e.g., haploidentical vs. matched sibling), concomitant immunosuppressants (e.g., MTX, MMF), and renal function. In HSCT, C0 is typically targeted at 200–400 ng/mL during the first month, with dose reductions guided by toxicity and chimerism status.

Applications in acquired bone marrow failure syndromes

Severe aplastic anemia (SAA)

The combination of CsA with anti-thymocyte globulin (ATG) constitutes the standard IST regimen for SAA, with efficacy approaching that of HSCT. European Blood and Marrow Transplantation (EBMT) guidelines recommend equine ATG at 40 mg/kg/day for 4 days combined with CsA at 5 mg/kg/day, targeting trough concentrations of 150–250 ng/mL [27]. Prospective cohort studies demonstrate that this regimen achieves overall response rates (ORR) of 60–80% and 5-year survival rates of 70–85% [28]. Notably, eltrombopag (EPAG) directly stimulates residual hematopoietic stem/progenitor cells (HSPCs) through the c-Mpl pathway, generating synergistic effects with CsA-mediated immunosuppression. A 2022 European multicenter randomized trial demonstrated that the ATG + CsA+EPAG arm achieved a 6-month ORR of 68%, significantly higher than the 41% observed with standard therapy (p < 0.001), without increased clonal evolution rates [29, 30]. This suggests that the immunosuppressive effects of CsA and the hematopoietic stimulation by TPO-RAs produce genuine synergy rather than simple additive effects.

Key advances lie in individualized therapy: recent investigations have identified that patients harboring STAT3 mutations exhibit poor responses to CsA, necessitating early consideration of HSCT [31]. Conversely, patients with telomere lengths > 5 kb demonstrate significantly higher IST response rates [32].

Non-severe AA (NSAA) and chronic AA

For transfusion-dependent NSAA, CsA monotherapy (3–5 mg/kg/day) may serve as a first-line option. A Japanese Phase III trial (JSHCT-NSAA-17) demonstrated 58% ORR at 6 months with CsA treatment, with a median time to response of 11 weeks [33]. Recent data indicate that the cumulative incidence of MDS/AML following IST is approximately 10–15%, with genomic instability effects of glucocorticoids potentially contributing more significantly, while the independent impact of CsA on this risk requires validation through long-term cohort studies [34–36]. Table 3 presents a head-to-head comparison of CsA and glucocorticoids in the treatment of acquired AA.

Table 3.

Head-to-Head Comparison of CsA and Glucocorticoids in Acquired AA Treatment

Comparison Dimension	CsA	Glucocorticoids (GCs)	Clinical Implications
Mechanism of Action	CaN-NFAT pathway inhibition only, preserving partial immune surveillance	Broad genomic/non-genomic suppression, inducing HSPC apoptosis	CsA causes less direct HSPC damage
Time to Onset	2–4 weeks	3–7 days	GCs used for rapid control of severe bleeding
Long-term Survival	ATG + CsA 5-year OS 70–85%	GCs monotherapy 5-year OS < 40%	CsA is the core contributor to IST efficacy
Clonal Evolution Risk	MDS/AML 10–15% (ATG-associated)	Potentially higher (genomic instability)	CsA relatively safer
Immunotherapy Compatibility	Low-dose compatible with PD-1 inhibitors	Significantly impairs ICI efficacy	CsA preferred in ICI-containing regimens
Primary Toxicities	Nephrotoxicity, hypertension (reversible)	Osteoporosis, diabetes, HPA axis suppression	Different comorbidity profiles drive preference

Note: AA = Aplastic Anemia; IST = Immunosuppressive Therapy; HSPC = Hematopoietic Stem/Progenitor Cell; ICIs = Immune Checkpoint Inhibitors; HPA = Hypothalamic-Pituitary-Adrenal axis.

Controversies and positioning in myelodysplastic syndromes (MDS)

The application of CsA in MDS remains among the most controversial areas in hematology. Early small-sample studies suggested that CsA could improve transfusion dependency in low-risk MDS patients who were HLA-DR15 positive or displayed hypoplastic bone marrow [37, 38]. However, the MDS-004 randomized controlled trial (RCTs) (n = 237) demonstrated that CsA monotherapy failed to improve erythroid or platelet responses [39, 40]. Notably, this trial enrolled patients without TCR repertoire screening, potentially diluting efficacy signals in immune-driven MDS subpopulations. This highlights that CsA indications should not rely on morphological diagnosis alone but rather on immunopathological stratification.

Nevertheless, recent large-scale cohort studies have refined our understanding of potential beneficiary populations for IST. A multi-center international cohort analysis of 207 MDS patients revealed that hypocellular bone marrow (cellularity < 20%) was the strongest predictor of achieving transfusion independence, with horse ATG plus CsA demonstrating superior efficacy compared to rabbit ATG or ATG without CsA [40], among low-risk MDS patients with STAT3 mutations and abnormal TCR Vβ repertoires, CsA at 3 mg/kg/day increased transfusion independence rates to 43% (compared to 8% in controls) [41]. Mechanistically, patients with immune-mediated MDS pathogenesis exhibit distinct T-cell dysregulation patterns. Kordasti et al. demonstrated that low-risk MDS is characterized by elevated Th17/Treg ratios in the bone marrow microenvironment, with increased Th17 cells correlating with apoptotic indices and proinflammatory cytokine profiles [42]. These immunological signatures, particularly the Th17/Treg imbalance, may identify patients likely to respond to CsA-mediated suppression of pathological Th17 responses [43]. The integration of such biomarkers with traditional clinical parameters (HLA-DR15 status, PNH clone positivity, and bone marrow cellularity) opens new avenues for precision immunotherapy in MDS, though prospective biomarker-stratified trials are still needed to validate these predictive models [44].

Prevention and treatment of GVHD

Prophylactic applications

The immunomodulatory role of CsA in HSCT is context-dependent, varying according to donor type, conditioning regimen, and concomitant immunosuppressive agents [45]. In HLA-matched sibling donor transplants, CsA primarily functions to inhibit early T-cell activation post-transplant, thereby reducing the incidence of aGVHD without abrogating the graft-versus-leukemia (GVL) effect [46]. In contrast, in haploidentical HSCT, where greater histoincompatibility necessitates intensified immunosuppression, CsA is commonly combined with agents such as mycophenolate mofetil, ATG, or post-transplant cyclophosphamide (PTCy) to target both donor-derived alloreactive T cells and recipient antigen-presenting cells [47, 48]. These multimodal strategies aim to balance effective GVHD prevention with adequate immune reconstitution and infection control.

The combination of CsA and short-course methotrexate remains one of the cornerstone regimens for GVHD prophylaxis following allogeneic HSCT [49]. The U.S. FDA recommends initiating CsA at 1–3 mg/kg/day via continuous intravenous infusion, followed by conversion to oral administration at 3–12 mg/kg/day in divided doses, with target trough concentrations of 200–400 ng/mL. Real-world data from Chinese multicenter studies illustrate the variability in outcomes based on transplant context. The CBMGT-2012 study reported incidences of aGVHD and cGVHD at 36% and 45%, respectively, with CsA demonstrating comparable efficacy to tacrolimus but significantly lower nephrotoxicity (18% vs. 29%, p = 0.02) [50]. In HLA-identical sibling transplants, aGVHD incidence was 23.2%, with a higher cGVHD rate of 67.4% [51]. Prospective randomized trials have confirmed that CsA/MTX regimens offer GVHD prevention comparable to tacrolimus-based protocols, with adjunctive ATG providing particular benefit in reducing severe aGVHD (grades III–IV) from 13% to 4% (p = 0.04) [52]. Pharmacovigilance analyses encompassing over 50,000 real-world cases reveal distinct toxicity profiles between CNIs: tacrolimus was more frequently associated with kidney injury requiring hospitalization (44.5% vs. 34.4%), whereas CsA exhibited higher associated mortality (17.1% vs. 12.7%) [53].

Contemporary optimization strategies have focused on novel immunosuppressive combinations. For haploidentical transplantation, regimens incorporating CsA with MMF and ATG have achieved grade III–IV aGVHD rates as low as 12%. Systematic reviews further support the use of rabbit ATG combined with PTCy, CsA, and MMF, reporting reductions in severe aGVHD to 5–13.5% [54] Pharmacokinetic optimization through Bayesian limited sampling strategies (B-LSS) for area-under-the-curve (AUC) monitoring enables precise dose individualization using concentrations at 0.5 and 6 h post-dose, demonstrating high correlation with total AUC (R²=0.86 for oral administration) and reducing dose adjustment frequency by approximately 30% compared to conventional trough concentration monitoring alone [55, 56].

Therapeutic applications

For steroid-refractory aGVHD (SR-aGVHD), CsA serves as salvage therapy. Phase II studies demonstrate that the CsA + ruxolitinib combination achieves ORR of 68% in this subgroup, with 6-month survival of 52% [57]. Ruxolitinib blocks Th1/Th17 differentiation through JAK-STAT pathway inhibition, creating dual signal interception with CsA-mediated CaN-NFAT axis suppression [58]. The REACH1 study demonstrated 12-month survival of 64% with this combination in SR-aGVHD, suggesting that multi-target immunosuppression can overcome single-pathway resistance [59]. In cGVHD treatment, CsA ophthalmic emulsion (0.05%) for ocular cGVHD significantly improves Schirmer test results (p < 0.01).

Autoimmune hematological diseases

Immune thrombocytopenia (ITP)

Thrombopoietin receptor agonists (TPO-RAs), such as eltrombopag and romiplostim, are currently established as the standard second-line therapy for patients with glucocorticoid-resistant or relapsed ITP [60, 61]. In this context, CsA is generally positioned as a third-line or alternative immunosuppressive agent, particularly when TPO-RAs are contraindicated, ineffective, or not tolerated. A meta-analysis incorporating seven studies (n = 418) demonstrated that CsA-based regimens achieved improved overall response rates in both children and adults with ITP, establishing its efficacy as a steroid-sparing agent [62]. In steroid-refractory patients specifically, CsA at 3–5 mg/kg/day for 3 months achieved sustained response rates of approximately 37–47%, with median time to response of 6 weeks [63]. This sustained response rate is comparable to initial GCs response rates, yet CsA lacks the osteoporosis and metabolic disorder risks associated with long-term GCs use, making it suitable for long-term maintenance in diabetic or elderly patients.

The 2023 guidelines from the Spanish ITP Working Group recommend immunosuppressants, including CsA, as viable options for moderate disease, particularly when TPO-RAs are contraindicated or insufficient [64]. For chronic ITP requiring long-term maintenance, CsA may be combined with low-dose prednisone (10–20 mg/day) to reduce the need for high-dose monotherapy, with combination therapy achieving response rates of up to 60% compared to 50% with CsA alone [25]. Notably, unlike TPO-RAs which have been associated with increased risk of thrombosis due to platelet count fluctuations, CsA does not significantly increase thrombotic risk, offering a safety advantage in patients with cardiovascular comorbidities.

Autoimmune Hemolytic Anemia (AIHA)

In warm-antibody AIHA, CsA as third-line therapy (for rituximab-failure patients) achieves ORR of 60–70% [65]. This CNIs exerts its immunosuppressive effects by blocking IL-2 production and T-cell activation, thereby attenuating the autoreactive B-cell response responsible for erythrocyte destruction [66]. However, the clinical utility of CsA in warm AIHA is often tempered by its nephrotoxicity, hypertension, and the requirement for TDM to maintain trough levels between 100 and 200 ng/mL [67]. In cold agglutinin disease (CAD), a distinct disorder driven by IgM autoantibodies activating the classical complement pathway, CsA can suppress complement-mediated hemolysis through downregulation of complement-regulatory protein expression on immune cells [68]. The drug is frequently combined with rituximab to achieve synergistic immunomodulation, resulting in 78% transfusion independence at 12 months [69]. Notably, CsA appears particularly effective in CAD with detectable bone marrow clonal lymphoproliferative disorders, where it may target both the underlying B-cell clone and the pathogenic autoantibody production [70]. Recent prospective data suggest that early introduction of CsA in rituximab-refractory CAD may prevent chronic complement exhaustion and reduce the risk of thromboembolic complications associated with persistent hemolysis [71]. Nevertheless, the optimal sequencing of CsA relative to novel complement inhibitors such as sutimlimab remains undefined, necessitating individualized treatment algorithms based on complement C4 levels, transfusion burden, and thrombotic risk profiles [72].

Exploratory applications in hematological malignancies

Acute myeloid leukemia (AML)

CsA application in AML primarily relies on its multidrug resistance (MDR) reversal properties. P-gp-mediated resistance constitutes a critical mechanism of AML relapse, with CsA serving as a P-gp inhibitor to restore chemotherapy sensitivity. The CALGB-19,808 trial demonstrated that DA regimen (daunorubicin and cytarabine) plus CsA (16 mg/kg/day) increased complete remission (CR) rates from 35% to 52% in P-gp-positive AML (p = 0.03) [73]. However, the subsequent SWOG-9126 study was prematurely terminated due to CsA-related toxicity, indicating the need for dose optimization [74]. Modern strategies have shifted toward low-dose CsA combined with targeted agents: preclinical studies demonstrate that CsA at 2 µM concentrations significantly enhances venetoclax-mediated clearance of AML stem cells, involving inhibition of mitophagy [75].

Lymphoma and multiple myeloma

Unlike the central position of glucocorticoids in lymphoma treatment, CsA applications remain limited. However, in angioimmunoblastic T-cell lymphoma (AITL), which derives from Tfh cells and is characterized by unique immune dysregulation [76], CsA can ameliorate immune dysfunction through suppression of Tfh cell function and inhibition of aberrant T-cell activation [77]. In a prospective study of twelve patients with relapsed/refractory AITL treated with CsA in combination with prednisolone and high-dose intravenous immunoglobulin, an ORR of 75% was achieved, including CR rates of 33% and median progression-free survival of 25.5 months [78]. A retrospective literature review of 26 AITL patients receiving CsA monotherapy similarly demonstrated an ORR of 86% in the second-line or later setting [78]. In multiple myeloma (MM), CsA primarily serves supportive roles. As a MDR modulator, CsA can circumvent clinical resistance to chemotherapy by blocking P-glycoprotein function, enabling cytotoxic drugs to eliminate resistant myeloma cells [79].

Additionally, CsA has been explored for its potential to induce an autologous graft-versus-host-like immune response following high-dose chemotherapy and autologous stem cell transplantation, aimed at enhancing graft-versus-tumor effects in multiple myeloma [80]. Notably, unlike glucocorticoids, which can antagonize the immunomodulatory effects of immunomodulatory drugs (IMiDs) such as lenalidomide by potentially impairing T-cell activation and cytokine production [81], CsA does not interfere with lenalidomide’s immunomodulatory mechanisms, representing a pharmacological advantage in combination strategies [82].

Therapeutic drug monitoring and adverse event management

Therapeutic drug monitoring (TDM)

Given its narrow therapeutic index and substantial inter-individual pharmacokinetic variability, largely attributable to CYP3A4/5 polymorphisms, drug-drug interactions, and clinical factors such as organ dysfunction, TDM remains essential for optimizing CsA therapy [83]. Two primary monitoring strategies are employed: trough level monitoring (C0) and 2-hour post-dose monitoring (C2). C0, measured immediately before the next dose, is widely used for convenience but may not fully reflect overall drug exposure. In contrast, C2, typically corresponding to the peak concentration for CsA microemulsion formulations, correlates more strongly with the area under the concentration-time curve from 0 to 4 h (AUC0–4), offering a more reliable surrogate for absorption-phase exposure [84].

Notably, racial disparities in transplant outcomes persist despite standardized immunosuppression; Black recipients exhibit higher rejection rates independent of drug monitoring strategy, underscoring the need for individualized approaches [85]. While CNIs including CsA may reduce peripheral Treg frequency, recent evidence suggests that optimized C2 monitoring can achieve adequate immunosuppression while potentially preserving immune regulatory function, particularly when lower target ranges (e.g., target C0 of 100–200 ng/mL for AA or C2 of 400–600 ng/mL) are employed in specific patient populations to balance efficacy with toxicity [86–88]. This C2 range is commonly used in transplant settings to optimize drug exposure while minimizing nephrotoxicity and other adverse effects.

The timing and frequency of TDM vary by clinical context. In patients undergoing HSCT, CsA levels are typically monitored twice weekly during the first 2–4 weeks post-transplant, then weekly thereafter until stable concentrations are achieved. Dose adjustments are guided by both efficacy and toxicity, with target C0 levels of 200–400 ng/mL for GVHD prophylaxis. For steroid-refractory aGVHD, higher targets (C0: 250–400 ng/mL) may be considered, though with increased nephrotoxicity risk [89]. In contrast, for AA and low-risk MDS, lower targets (C0: 100–250 ng/mL) are recommended to balance immunosuppression with hematopoietic recovery [90,91]. In the pre-transplant setting, CsA is not typically administered, but for patients receiving CsA as part of bridging therapy prior to CAR-T infusion, discontinuation at least 2 weeks before lymphodepletion is advised to avoid impairing CAR-T cell expansion [92]. Post-transplant, CsA is initiated 1–3 days before HSCT or on day − 1, with levels monitored closely to ensure therapeutic coverage during engraftment.

Major toxicities and interventions

Nephrotoxicity represents the dose-limiting toxicity of CsA, occurring in 30–50% of patients, associated with endothelin-1 upregulation and TGF-β-mediated renal interstitial fibrosis [93]. Table 4 presents the toxicity stratified management and intervention thresholds for long-term CsA use. Intervention strategies include:

Table 4.

Stratified Toxicity Management and Intervention Thresholds for Long-Term CsA Use

Toxicity Type	Early Markers (1–3 months)	Intermediate Markers (3–12 months)	Interventions	Conversion Therapies
Nephrotoxicity	Serum creatinine ↑ >10%	eGFR↓<60 mL/min	Amlodipine, dose reduction to C2 < 200	Sirolimus, belatacept
Hypertension	Systolic BP > 140 mmHg	Requiring ≥ 2 antihypertensive agents	ACEI/ARB preferred	Conversion to non-nephrotoxic regimens
Neurotoxicity	Hand tremor, headache	Progressive cognitive decline	Dose reduction, magnesium supplementation	Tacrolimus (risk-benefit assessment)
Infection Risk	No specific markers	Severe infections ≥ 2/year	Immunoglobulin replacement	Conversion to MMF monotherapy
Drug Interactions	CYP3A4 inhibitor co-administration	Blood concentration fluctuation > 30%	Genotype-guided dose adjustment	Avoid co-administration with voriconazole

Note: eGFR = estimated Glomerular Filtration Rate; ACEI = Angiotensin-Converting Enzyme Inhibitor; ARB = Angiotensin Receptor Blocker; MMF = Mycophenolate Mofetil; CYP3A4 = Cytochrome P450 3A4.

Maintaining trough concentrations < 250 ng/mL for AA

This safety threshold is supported by clinical trial protocols indicating that dose reduction should be considered when C0 exceeds 250 ng/mL to mitigate nephrotoxicity risk [94]. Pediatric dosing guidelines similarly recommend target ranges of 200–400 ng/mL for AA, with lower targets indicated when nephrotoxicity is confirmed [95]. These thresholds balance immunosuppressive efficacy against renal injury, as CsA-associated nephrotoxicity becomes more pronounced at supratherapeutic levels [96].

Maintaining trough concentrations < 400 ng/mL for GVHD

This upper limit aligns with standard prophylactic regimens targeting 200–400 ng/mL in allogeneic HSCT recipients [15]. Evidence from myeloablative transplant recipients demonstrates that maintaining mean CsA trough concentrations ≥ 250 ng/mL during days 15–28 post-transplant significantly reduces grade II-IV aGVHD incidence (31.5% vs. 18.8%; OR 1.97), while levels exceeding 400 ng/mL increase nephrotoxicity without additional GVHD protection [9, 15].

Co-administration of calcium channel blockers (amlodipine) to reduce renal vasoconstriction

Calcium channel blockers (CCBs), such as amlodipine, counteract CsA-induced renal vasoconstriction by blocking L-type calcium channels on vascular smooth muscle cells, leading to afferent arteriolar dilation and increased renal plasma flow. This vasodilatory effect directly opposes the vasoconstrictive actions of endothelin-1 and thromboxane A2, which are upregulated by CNIs [97].

Conversion to sirolimus or belatacept when necessary

In patients with CsA-refractory nephrotoxicity (eGFR decline > 30% from baseline or biopsy-proven CNI nephrotoxicity), conversion to sirolimus, which is an mTOR inhibitor with a distinct mechanism targeting the IL-2 receptor signaling pathway rather than calcineurin, has been associated with significant renal function recovery in HSCT recipients, without increased GVHD relapse rates [98]. Similarly, belatacept, a selective costimulation blocker, has demonstrated superior renal preservation compared to continued CNI-based regimens in solid organ transplantation, though its application in hematological populations remains investigational [99].

Combination strategies with novel therapeutics

Immune checkpoint inhibitors (ICIs)

The interplay between CsA and ICIs represents a therapeutic paradox that has garnered considerable clinical interest. While conventional wisdom suggests that CsA-mediated suppression of T-cell activation might attenuate the efficacy of anti-PD-1 therapy, emerging evidence paints a more nuanced picture. Preclinical investigations have demonstrated that low-dose CsA (1–2 mg/kg) exhibits selective immunomodulatory properties, effectively dampening pathological Th17 responses while sparing CD8 + cytotoxic T-cell-mediated anti-tumor immunity [100]. This differential sensitivity appears rooted in the distinct signaling requirements of these T-cell subsets; Th17 cells display marked susceptibility to calcineurin inhibition, whereas CD8 + effector functions remain relatively preserved [101].

Translating these findings to the clinical area, retrospective analyses of patients with hematologic malignancies have provided reassuring safety signals. Notably, in a cohort of patients who relapsed after HSCT and subsequently received nivolumab as salvage therapy, concomitant administration of low-dose CsA for GVHD prophylaxis was associated with preserved anti-tumor responses and a reduced incidence of severe immune-related adverse events [102]. These patients, predominantly with lymphoid malignancies, demonstrated that CsA did not compromise the efficacy of PD-1 blockade while potentially mitigating immune-mediated toxicities. The overall response rate remained robust and comparable between groups (51% versus 58%, p = 0.31), suggesting that CsA co-treatment did not substantially compromise ICI efficacy. Equally compelling was the observed improvement in toxicity profiles: the incidence of immune-mediated colitis dropped significantly from 28% to 12% among patients receiving CsA, highlighting its potential protective role against ICI-related gastrointestinal toxicities [103, 104]. This dual benefit—maintained anti-tumor activity coupled with reduced immune-related adverse events—positions low-dose CsA as a promising candidate for concurrent immunosuppression in carefully selected patients undergoing ICI therapy post-allogeneic transplantation [105].

Chimeric antigen receptor T-Cell (CAR-T) therapy

CsA plays a limited role in the management of CAR-T-associated toxicities. By inhibiting calcineurin, CsA suppresses T-cell activation and proliferation through the calcineurin-NFAT pathway, which may result in permanent impairment of CAR-T cell persistence and compromise long-term anti-tumor efficacy [8]. It should be noted that in CAR-T bridging therapy, the calcineurin inhibitory effects of CsA may permanently impair CAR-T cell persistence, whereas tocilizumab does not affect T-cell function. Therefore, CsA should be discontinued 2 weeks prior to CAR-T infusion [106]. In contrast, tocilizumab, an IL-6 receptor antagonist, remains the established first-line therapy for cytokine release syndrome (CRS) following CAR-T cell infusion, demonstrating rapid reversal of CRS symptoms without compromising CAR-T cell function or anti-tumor responses [107]. Importantly, for patients who receive CsA as bridging therapy prior to CAR-T cell infusion, no washout period is required before lymphodepleting chemotherapy, as the suppressive effects of CsA on CAR-T cell function appear to be reversible upon discontinuation [108].

Combination with targeted agents

CsA combined with JAK inhibitors (such as ruxolitinib) demonstrates synergistic effects: CsA inhibits the CaN-NFAT axis while ruxolitinib blocks the JAK-STAT pathway, achieving dual suppression of Th1/Th17 differentiation [109, 110]. Preclinical studies have demonstrated that this dual inhibition significantly improves hematologic parameters and immune recovery. The JAK-STAT signaling pathway plays a central role in acute GvHD pathogenesis, and ruxolitinib has shown superiority over conventional second-line therapies. In SR-aGVHD, this combination improved 12-month survival to 64% [111]. In the pivotal REACH1 trial, ruxolitinib combined with corticosteroids showed durable responses and encouraging survival rates compared with historical data in patients with steroid-refractory disease. Subsequent real-world experiences have confirmed the efficacy of ruxolitinib therapy in this challenging population.

The combination of GCs with CsA for rapid inflammation control is rational: the rapid non-genomic effects of GCs can acutely suppress inflammatory responses, while CsA’s immunomodulatory effects sustain remission [112]. This sequential approach aligns with current guidelines where corticosteroids serve as first-line agents for acute GVHD treatment. Combination therapies using CNIs with post-transplant cyclophosphamide or mTOR inhibitors have demonstrated improved GVHD control with reduced late toxicities [113,114]. However, vigilance is required regarding synergistic infection risk; a “GCs rapid taper + CsA maintenance” sequential strategy is recommended [115]. This strategy mirrors approaches used in other immunosuppressive protocols where minimizing long-term corticosteroid exposure is prioritized.

For patients requiring long-term immunosuppression, CsA monotherapy or combination with non-steroidal immunosuppressants should be prioritized to avoid cumulative toxicities associated with GCs, including HPA axis suppression, osteoporosis, and metabolic disorders [116, 117]. Long-term corticosteroid use is associated with significant endocrine complications, including hypothalamic-pituitary-adrenal axis suppression and osteoporosis, affecting up to 0.8% of patients with documented bone loss. The cumulative toxicities of prolonged corticosteroid therapy necessitate transition to CNI-based maintenance regimens. Furthermore, CNIs themselves require monitoring for metabolic and endocrine toxicities, including effects on bone health and gonadal function. Table 5 illustrates the synergistic mechanisms and evidence hierarchy of CsA co-administration with novel therapeutic agents.

Table 5.

Synergistic Mechanisms and Evidence Levels for CsA Combined with Novel Agents

Combination	Synergistic Mechanism	Indication	Key Studies	Precautions
CsA + EPAG	Immunosuppression + HSPC stimulation	IST-naïve SAA	NEJM 2017 [118]	No significant increased clonal evolution observed
CsA + Ruxolitinib	CaN-NFAT + JAK-STAT dual inhibitors	SR-aGVHD	Blood 2020 [119]	Increased infection risk
CsA + Venetoclax	P-gp inhibition + BCL-2 inhibition	Relapsed AML	Blood 2019 [120]	CsA dose reduction required to prevent nephrotoxicity
CsA + Nivolumab	Immune modulation preserving CD8 + function	Post-HSCT relapse	Blood 2017 [121]	Limited to low-dose CsA
CsA + Androgens	Immunosuppression + direct hematopoietic stimulation	Chronic AA	Hematology 2022 [122]	Better tolerated in female patients

Note: EPAG = Eltrombopag; HSPC = Hematopoietic Stem/Progenitor Cell; IST = Immunosuppressive Therapy; SAA = Severe Aplastic Anemia; SR-aGVHD = Steroid-Refractory Acute Graft-versus-Host Disease; AML = Acute Myeloid Leukemia; HSCT = Hematopoietic Stem Cell Transplantation; AA = Aplastic Anemia; RCT = Randomized Controlled Trial.

Future directions and conclusions

Current evidence supporting CsA applications in hematological disorders derives predominantly from retrospective studies or single-arm trials, with a paucity of large-scale randomized controlled trials validating its superiority over novel immunosuppressive agents. Furthermore, biomarker-guided individualized therapeutic strategies await prospective confirmation of their clinical value. Future research will focus on developing CsA nanoformulations to enhance oral bioavailability while reducing nephrotoxicity; exploring artificial intelligence-assisted TDM systems for real-time dose optimization; and conducting prospective randomized trials based on biomarkers including STAT3 mutations and TCR repertoire profiling to validate the clinical benefits of precision stratification strategies.

Author contributions

Zhengwei Tan and Jinyu Hu develop the protocol, collect the data and write the manuscript; Baodong Ye and Wenbin Liu revised the manuscript. All authors read and approved the final version of the manuscript.

Funding

The present study was supported by “Leading Geese” Research and Development Plan of Zhejiang Province (NO. 2024C03190), National Natural Science Foundation of China (NO. 82274273, 81774092).

Data availability

No datasets were generated or analysed during the current study.

Declarations

Ethical approval and consent to participate

Not applicable.

Consent for publication

Not applicable.

Competing interests

The authors declare no competing interests.