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Abstract

Background:

In haploidentical hematopoietic stem cell transplantation (haplo-HSCT), combining low-dose post-transplant cyclophosphamide (PTCy) with low-dose anti-thymocyte globulin (ATG) is increasingly recognized as a promising approach for graft-versus-host disease (GVHD) prevention.

Methods:

This study evaluated 33 patients undergoing haplo-HSCT for hematological disorders, divided into two groups: low-dose ATG/PTCy (n = 17) and PTCy-only (n = 16).

Results:

The incidence of grades I–II acute GVHD (aGVHD) was 11.8% in the low-dose ATG/PTCy group compared to 31.3% in the PTCy-only group (p = 0.42). No cases of severe aGVHD (grades III–IV) were reported in either cohort. Moderate chronic GVHD (cGVHD) occurred less frequently in the ATG/PTCy group (28.6%) compared to the PTCy group (100%, p = 0.028). Severe cGVHD was absent in both groups. Non-relapse mortality (NRM) was significantly lower in the ATG/PTCy group compared to the PTCy-only group (17.6% vs 56.3%, p = 0.021). One year overall survival and disease-free survival rates were at 70.6% and 64.7% for ATG/PTCy cohort, versus 56.3% and 50.0% for PTCy-only group. Cytomegalovirus reactivation and relapse were comparable between the groups.

Conclusion:

The combination of low-dose ATG and PTCy appears to significantly reduce moderate cGVHD and NRM in haplo-HSCT compared to PTCy alone. To the best of our knowledge, this is the first study directly comparing these two regimens.

Plain language summary

Low-dose ATG/post-transplant cyclophosphamide for GVHD proflaxis

In haplo-HSCT, combining low-dose post-transplant cyclophosphamide (PTCy) with low-dose anti-thymocyte globulin (ATG) is increasingly recognized as a promising approach for graft-versus-host disease (GVHD) prevention. ATG and PTCy are two widely used in vivo TCD strategies in haplo-HSCT. However, there is a lack of consensus on the optimal strategy for in vivo TCD. To the best of our knowledge, this study is the first study directly comparing PTCy alone versus low-dose ATG/PTCy combination.

Keywords

anti-thymocyte globulin graft-versus-host disease haploidentical low-dose post-transplant cyclophosphamide

Introduction

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a potentially curative treatment for hematological malignancies and some non-malignant conditions. However, its success is limited by complications such as graft-versus-host disease (GVHD), particularly in haploidentical HSCT (haplo-HSCT), where human leukocyte antigen disparity increases the risk of GVHD.^1,2

Among GVHD prophylaxis strategies, in vivo T-cell depletion with anti-thymocyte globulin (ATG) or post-transplant cyclophosphamide (PTCy) has become standard practice.^3–5 Both agents have shown efficacy in reducing GVHD rates, yet each presents limitations. While PTCy effectively eliminates alloreactive T cells, its protective effects may be reduced in peripheral blood stem cell transplants.⁶ ATG, on the other hand, can delay immune reconstitution and increase infection risk.⁷

To improve outcomes, recent studies have explored combining ATG and PTCy at lower doses. This approach aims to balance GVHD prevention with preserved immune recovery. For example, Herzog et al.⁸ highlighted that prophylaxis strategies must consider conditioning regimens and graft sources to optimize outcomes, while Asensi et al. reported improved response to GVHD therapy in PTCy-based protocols.⁹

Despite promising results, data on the efficacy and safety of combining low-dose ATG with PTCy remain limited. This study aims to compare clinical outcomes in haplo-HSCT recipients who received GVHD prophylaxis with either PTCy alone or in combination with low-dose ATG. By evaluating GVHD incidence, survival outcomes, and infection risks, we aim to contribute to the refinement of GVHD prophylaxis strategies in haplo-HSCT.

This study compares the transplantation outcomes of haplo-HSCT patients receiving GVHD prophylaxis with PTCy alone versus those treated with the low-dose ATG/PTCy combination.

Materials and methods

A total of 33 patients who were treated with haplo-HSCT at our center were included in this retrospective study between January 2015 and January 2024. Inclusion criteria were: (i) patients aged ⩾18 years, (ii) diagnosed with hematologic malignancy or relevant non-malignant disease, and (iii) undergoing haploidentical HSCT at our institution within the defined study period. Exclusion criteria included: (i) cord blood transplantation, (ii) matched unrelated donor HSCT, and (iii) lack of follow-up data or missing outcome documentation. All patients who presented to the clinic within the defined study period were included in the study. The patients were classified into two groups: one treated with a combination regimen of low-dose ATG and PTCy, and the other treated with solely on PTCy. Table 1 provides a detailed summary of the participants’ clinical profiles, including their underlying conditions and transplantation processes.

Table 1.

Patient, disease, and transplantation characteristics.

Transplantation characteristics	PTCy group (n = 16)	Low-dose ATG/PTCy group (n = 17)	p
Age, years, median (min.–max.)	43.5 (20–67)	43 (19–69)	0.977^a
Sex (female), n (%)	7 (43.8)	5 (29.4)	0.392^b
Disease type, n (%)
AML	7 (43.8)	13 (76.5)	—
ALL	5 (31.3)	1 (5.9)
Lymphoma	2 (12.5)	0
Myelofibrosis	0	1 (5.9)
Other	0	1 (5.9)
Conditioning regimen, n (%)
MAC	12 (75.0)	10 (58.8)	0.325^b
RIC	4 (25)	7 (41.2)
Blood type matching (matched), n (%)	10 (62.5)	11 (64.7)	0.895^b
Prior transplantation status (yes), n (%)	1 (86.3)	4 (23.5)	0.335^c
CD34 cells (10⁶/kg), median (min.–max.)	7.43 (2.30–10.10)	8.00 (5.36–13.84)	0.191^d
Disease status at transplantation
Complete response, n (%)	16 (100.0)	16 (94.1)	1.000^d
Partial response, n (%)	0	1 (5.9)

Independent samples t test.

Pearson Chi-Square test.

Mann–Whitney U test.

Fisher’s Exact test.

ATG, anti-thymocyte globulin; MAC, myeloablative conditioning; PTCy, post-transplant cyclophosphamide; RIC, reduced intensity conditioning.

Ethical approval of the study was obtained by the Erciyes University Faculty of Medicine Ethics Committee (Date: 21 August 2024, Decision Number: 2024/136).

GVHD prophylaxis and conditioning regimen

In the low-dose ATG/PTCy group, ATG was used at 2.5 mg/kg/day on days −2 and −1, for a total of 5 mg/kg. PTCy was used as a one-dose of 50 mg/kg on day +3. Cyclosporine A was initiated on day +4, and Mycophenolate mofetil (MMF) was administered orally at 1 g three times daily from day +5 to day +34, provided no acute GVHD (aGVHD) developed.¹⁰

In the PTCy group, PTCy was used at 50 mg/kg/day on days +3 and +4, with mesna immunosuppressive therapy included cyclosporine A at 3 mg/kg/day starting on day +5 and MMF at 15 mg/kg three times daily, initiated on day +5 and continued as needed.^11–13

Conditioning regimens were tailored to individual patient profiles. Reduced-intensity conditioning was selected for patients aged 55 years or older or those with an HCT-comorbidity index score >2. Myeloablative conditioning was used for all other patients.¹⁴

Supportive care

All participants were administered granulocyte colony-stimulating factor (G-CSF) starting on day +5 at a dose of 5 µg/kg/day, which was continued until neutrophil recovery. Prophylaxis with levofloxacin and valacyclovir was initiated alongside the conditioning regimen and maintained until neutrophil counts normalized. Antifungal prophylaxis began at the start of conditioning and was continued for a minimum of 3 months post-transplant.

Cytomegalovirus (CMV) DNA levels in serum and Epstein-Barr virus (EBV) DNA in whole blood were monitored weekly using quantitative polymerase chain reaction. This monitoring continued through at least day +100 post-transplant.

Letermovir prophylaxis was not employed in this cohort due to a lack of access at the time of treatment.

Definitions

Neutrophil engraftment was characterized by maintaining an absolute neutrophil count of ⩾0.5 × 10⁹/L for a minimum of 3 consecutive days without the aid of G-CSF following transplantation. Platelet engraftment was identified by achieving a platelet count of ⩾20 × 10⁹/L for at least 7 consecutive days without platelet transfusions. Full donor chimerism was described as having ⩾95% donor-derived T cells in peripheral blood samples.¹⁵ Graft failure was categorized into primary graft failure (failure to achieve neutrophil engraftment by day 28 following transplantation) and secondary graft failure (loss of donor chimerism after successful engraftment without evidence of disease relapse).^16,17

aGVHD was diagnosed and graded based on the modified Glucksberg criteria, whereas chronic GVHD (cGVHD) was assessed using the 2014 National Institutes of Health consensus guidelines.¹⁸ Relapse was defined by the presence of ⩾5% blasts in bone marrow or the recurrence of the original disease. Non-relapse mortality (NRM) was defined as any death not resulting from disease relapse. Overall survival (OS) was determined from the date of stem cell infusion to death or final follow-up. Disease-free survival (DFS) was assessed as the period from transplantation to survival without signs of relapse or disease progression.¹⁹

The study evaluated a variety of clinical outcomes, including engraftment rates, the frequency and severity of aGVHD and cGVHD, incidences of viral reactivation (CMV and EBV), infections, OS, DFS, relapse, and NRM.

Statistical analysis

All statistical analyses were performed using IBM SPSS Statistics version 25.0 (IBM Corp., Armonk, NY, USA). Categorical variables were described as frequencies and percentages, while continuous variables were presented as medians along with their ranges (minimum to maximum). Group comparisons for categorical data were performed using either the chi-square test or Fisher’s exact test, depending on the context.

The Shapiro–Wilk test was employed to determine the normality of continuous variables. For datasets following a normal distribution, the independent samples t-test was applied, whereas the Mann–Whitney U test was used for non-normally distributed data. Kaplan–Meier methodology was utilized for survival analysis, and the log-rank test was employed to evaluate differences in survival curves. A p-value below 0.05 was considered statistically significant.

Results

The study encompassed 33 participants, comprising 17 individuals in the low-dose ATG/PTCy group and 16 in the PTCy-only group. The median age of the ATG/PTCy group was 43 years (range: 19–69 years), while the median age in the PTCy group was 43.5 years (range: 20–67 years), showing no significant difference (p = 0.97). Gender distribution showed that 29.4% of patients in the ATG/PTCy group were female and 70.6% were male, compared to 43.8% female and 56.2% male in the PTCy group (p = 0.39).

No significant statistical differences were identified between the two groups in terms of age, gender, conditioning regimen, blood type compatibility, CD34⁺ cell dose administered, or disease status (refer to Table 1). Acute myeloid leukemia was the most frequent diagnosis, comprising 76.5% of cases in the ATG/PTCy group and 43.8% in the PTCy group.

In the ATG/PTCy group, 64.7% of patients (11 individuals) had ABO-matched donors, compared to 62.5% (10 individuals) in the PTCy group. One patient in the ATG/PTCy group had previously undergone autologous HSCT, and three had a history of allo-HSCT. In the PTCy group, one patient had received an earlier autologous HSCT.

All patients and their donors were CMV IgG positive, indicating 100% seropositivity for CMV in the study population.

Engraftment and chimerism

In the low-dose ATG/PTCy cohort, the median time to platelet engraftment was 13 days (range: 9–35 days), while neutrophil engraftment was achieved at a median of 14 days (range: 10–27 days). Comparatively, the PTCy cohort showed a median platelet engraftment time of 15 days (range: 11–35 days) and a median neutrophil engraftment time of 16 days (range: 13–27 days). The differences in engraftment times for neutrophils (p = 0.10) and platelets (p = 0.29) were not statistically significant.

Neither group encountered primary or secondary graft failure. All patients in both groups successfully engrafted and demonstrated full donor chimerism in the whole blood compartment by day +30. Notably, chimerism levels were significantly higher in the low-dose ATG/PTCy group compared to the PTCy group (p = 0.049).

GVHD and other complications

Within 180 days post-transplant, aGVHD was observed in 11.8% of patients (2 out of 17) in the low-dose ATG/PTCy cohort, both presenting as grade I cases limited to skin involvement. These cases were effectively managed with topical steroids, with no cases involving the gut or liver and no reports of severe aGVHD (grades III–IV).

In the PTCy cohort, 31.3% of patients (5 out of 16) developed aGVHD during the same period. This included two grade I cases and three grade II cases. Like the ATG/PTCy group, no patients in this cohort experienced severe aGVHD (grades III–IV). The difference in the incidence of grade I–II aGVHD between the two groups was not statistically significant (p = 0.42).

At 1 year post-transplant, cGVHD was documented in 41.2% (7 out of 17) of the low-dose ATG/PTCy group, with moderate cGVHD occurring in 28.6% (2 out of 17). No severe cGVHD cases were recorded in this group. In the PTCy cohort, 31.3% of patients (5 out of 16) experienced cGVHD, with all cases classified as moderate (100%). While overall cGVHD rates were similar, the likelihood of moderate cGVHD was significantly lower in the low-dose ATG/PTCy group compared to the PTCy group (p = 0.028) (Table 2). Additionally, the overall incidence of chronic GVHD at 1 year was 41.2% in the low-dose ATG/PTCy group and 31.3% in the PTCy group, with no statistically significant difference (p = 0.554).

Table 2.

Clinical outcomes of patients in the two groups.

Clinical outcomes	PTCy group (n = 16)	Low-dose ATG/PTCy group (n = 17)	p
AGVHD within 180 days, n (%)	5 (31.3)	2 (11.8)	0.225^a
AGVHD n (%)
Grade I	2 (40.0)	2 (100.0)	0.429^a
Grade II	3 (60.0)	0
CGVHD at 1 year (yes), n (%)	5 (31.3)	7 (41.2)	0.554^b
CGVHD n (%)
Mild	0	5 (71.4)	0.028^a
Moderate	5 (100.0)	2 (28.6)
CGVHD type, n (%)
Skin	2 (40.0)	3 (42.9)
Liver	2 (40.0)	2 (28.6)	—
Skin + liver + intestine	1 (20.0)	1 (14.3)
Liver + intestine	0	1 (14.3)
Platelet engraftment, median (min.–max.)	15 (11–35)	13 (9–35)	0.295^c
Neutrophil engraftment, median (min.–max.)	16 (13–27)	14 (10–27)	0.105^c
Chimerism	98.50 (98.00–99.75)	99.00 (99.00–100.00)	0.049^a
Viral infections, n (%)	14 (87.5)	11 (64.7)	0.225^a
Polyomavirus BK virus, n (%)	5 (31.3)	5 (29.4)	1.000^a
CMV reactivation, n (%)	13 (81.3)	9 (52.9)	0.085^b
EBV, n (%)	0	1 (5.9)	1.000^a
Infection (non-viral), n (%)	13 (81.3)	10 (58.8)	0.259^a
Type of infection
Pnömonia + sepsis	9 (69.2)	4 (40)	—
Pnömonia	3 (23.1)	5 (50)
Sepsis	0	1 (10)
Sepsis + catheter infection	1 (7.7)	0
Pathogen of infection (non-viral), n (%)
Bacteria and fungus	8 (61.5)	3 (30.0)	—
Bacteria	4 (30.8)	6 (60.0)
Fungus	1 (7.7)	1 (10.0)
Relapse, n (%)	2 (12.5)	3 (17.6)	1.000^a
1-year relapse, n (%)	1 (6.3)	2 (11.8)	1.000
2-year relapse, n (%)	2 (12.5)	3 (17.6)	1.000
30-day mortality, n (%)	1 (6.3)	0	0.485^a
100-day mortality, n (%)	5 (31.3)	1 (5.9)	0.085^a
Mortality, n (%)	11 (68.8)	7 (41.2)	0.112^b
Main causes of deaths, n (%)
Bacterial infection	6 (54.5)	1 (14.3)	—
Relapse of primary disease	2 (18.2)	3 (42.9)
Organ failure	1 (9.1)	3 (42.9)
Cerebrovascular disease	1 (9.1)	0
Pnömonia	1 (9.1)	0
NRM, n (%)	9 (56.3)	3 (17.6)	0.021^b
1-year NRM, n (%)	7 (43.8)	2 (11.8)	0.057
2-year NRM, n (%)	9 (56.3)	3 (17.6)	0.021^b
Median follow-up months, median (min.–max.)	15 (1–84)	26.00 (2–118)	0.102^c
DFS, months, median (min.–max.)	12 (0.5–84)	26 (1.5–118)	0.110^c

Fisher’s Exact test.

Pearson Chi-Square test.

Mann–Whitney U test.

AGVHD, acute graft-versus-host disease; ATG, anti-thymocyte globulin; CGVHD, chronic graft-versus-host disease; CMV, cytomegalovirus; DFS, disease-free survival; EBV, Epstein-Barr virus; NRM, non-relapse mortality; PTCy, post-transplant cyclophosphamide.

Viral infections

Viral infections occurred in 64.7% (11 out of 17) of patients in the low-dose ATG/PTCy group and 87.5% (14 out of 16) in the PTCy group, though this was not statistically significant (p = 0.22) (Table 2). CMV reactivation was observed in 52.9% (9 out of 17) of the low-dose ATG/PTCy group compared to 81.3% (13 out of 16) in the PTCy group, suggesting a trend toward reduced reactivation in the ATG/PTCy cohort (p = 0.085). EBV reactivation was identified in 5.9% (1 out of 17) of patients in the ATG/PTCy group, while no cases were reported in the PTCy group. The incidence of BK virus infection was comparable between the groups, affecting 29.4% (5 of 17) in the low-dose ATG/PTCy cohort and 31.3% (5 of 16) in the PTCy cohort (p = 1.00).

Bacterial and fungal infections

In the low-dose ATG/PTCy group, bacterial infections occurred in 60.0% (6 out of 17) of patients, fungal infections in 10.0% (1 out of 17), and both bacterial and fungal infections in 30.0% (3 out of 17). Among PTCy group patients, bacterial infections were noted in 30.8% (4 out of 16), fungal infections in 7.7% (1 out of 16), and combined bacterial and fungal infections in 61.5% (8 out of 16). A detailed breakdown of these complications is provided in Table 2.

OS and DFS outcomes

The median follow-up duration was 26 months (range: 2–118 months) for patients in the low-dose ATG/PTCy group and 15 months (range: 1–84 months) for those in the PTCy group. In the low-dose ATG/PTCy group, OS was 70.6% (95% CI, 47.1%–88.2%) at 1 year, 52.9% (95% CI, 29.4%–76.5%) at 2 years, and 44.9% (95% CI, 23.3%–66.5%) at the last follow-up (Figure 1). For the PTCy group, OS was 56.3% (95% CI, 31.3%–81.3%) at 1 year, 37.5% (95% CI, 12.5%–67.5%) at 2 years, and 24.8% (95% CI, 10.0%–39.6%) at the last follow-up. Although the low-dose ATG/PTCy group demonstrated longer OS, a statistically significant difference was not found (p = 0.112) (Figure 1).

Figure 1.

Overall survival results of groups received low-dose ATG/PTCy or PTCy.

Similarly, DFS was longer in the low-dose ATG/PTCy group, with a median of 26 months (range: 1.5–118 months), compared to 12 months (range: 0.5–84 months) in the PTCy group; however, this difference did not reach statistical significance. In the low-dose ATG/PTCy group, DFS was 64.7% (95% CI, 41.2%–88.2%) at 1 year, 52.9% (95% CI, 29.4%–76.5%) at 2 years, and 43.8% (95% CI, 21.8%–65.8%) at the last follow-up (Figure 2). For the PTCy group, DFS was 50.0% (95% CI, 25.0%–75.0%) at 1 year, 31.3% (95% CI, 12.5%–56.3%) at 2 years, and 24.0% (95% CI, 9.2%–38.9%) at the last follow-up.

Figure 2.

Disease-free survival results of groups received low-dose ATG/PTCy or PTCy.

NRM, relapse, and overall mortality

Mortality occurred in 41.2% (7/17) of the low-dose ATG/PTCy group compared to 68.8% (11/16) in the PTCy group. Despite the higher mortality rate in the PTCy group, the difference was not statistically significant (p = 0.11). In the low-dose ATG/PTCy cohort, causes of death included bacterial infections (one patient), organ failure (three patients), and relapse of the underlying condition (three patients). In the PTCy cohort, deaths were attributed to bacterial infections (six patients), organ failure (one patient), disease relapse (two patients), pneumonia (one patient), and cerebrovascular disease (one patient). Neither group reported deaths related to acute or chronic GVHD.

The 30- and 100-day mortality rates were 0% (0/17), and 5.9% (1/17), respectively, in the low-dose ATG/PTCy group, versus 6.3% (1/16), and 31.3% (5/16), respectively in the PTCy group. Differences in mortality between the groups at day 30 (p = 0.48) and day 100 (p = 0.08) did not reach statistical significance.

At 1 year, relapse rates were 11.8% in the low-dose ATG/PTCy group and 6.3% in the PTCy group (p = 1.00), while at 2 years, they were 17.6% and 12.5%, respectively (p = 1.00).

NRM rates at 1 year were 11.8% for the low-dose ATG/PTCy group and 43.8% for the PTCy group (p = 0.057), and at 2 years, NRM was significantly lower in the ATG/PTCy group (17.6%) compared to the PTCy group (56.3%, p = 0.021) (Figure 3).

Figure 3.

Non-relapse mortality results of groups received low-dose ATG/PTCy or PTCy.

Discussion

In recent years, haplo-HSCT has emerged as the leading type of allo-HSCT for patients without matched donors globally.²⁰ Numerous modifications to PTCy-based protocols have been explored, including the integration of additional immunosuppressive agents. One prominent strategy is the combination of ATG with PTCy.^21,22 This combination may provide synergistic benefits in settings with a higher risk of GVHD, such as unrelated donor allo-HCT and haplo-HSCT.²³

In this study, the low-dose ATG/PTCy regimen was associated with a significantly reduced incidence of moderate cGVHD compared to the PTCy-only regimen (28.6% vs 100%, p = 0.028). Furthermore, the majority of cGVHD cases in the low-dose ATG/PTCy group were mild, and no severe cGVHD was observed. NRM was also markedly lower in the low-dose ATG/PTCy group (p = 0.021). To the best of our knowledge, this study is the first to directly compare the outcomes of a low-dose ATG/PTCy regimen with PTCy alone.

Li et al.²⁴ reported a 2-year moderate/severe cGVHD incidence of 18.08% in their long-term follow-up of low-dose ATG/PTCy recipients. Similarly, Xu et al.²⁵ noted a 1-year moderate/severe cGVHD incidence of 11.2%, which is lower than the rates observed in our study. But, these studies documented the coexistence of moderate and severe cGVHD rates. There were no cases of severe cGVHD in our cohort.

Luo et al.²⁶ conducted a meta-analysis revealing a significantly lower incidence of cGVHD in patients treated with low-dose ATG/PTCy compared to those receiving ATG alone. However, no meaningful difference was revealed when compared to PTCy-only regimens, which may be attributed to the 100 mg PTCy dose used in the combination regimen rather than a reduced dose.

For aGVHD, the meta-analysis by Luo et al.²⁶ revealed that haplo-HSCT with low-dose ATG/PTCy significantly reduced the incidence of grade II–IV aGVHD compared to ATG alone (p < 0.00001) or PTCy alone (p = 0.005). Other studies have reported grade II–IV and grade III–IV aGVHD ranging from 13.46% to 17.7% and 3% to 6.8%, respectively.^24,25

In our cohort, although the difference in aGVHD incidence between the low-dose ATG/PTCy and PTCy groups was not statistically significant, the low-dose ATG/PTCy group exhibited a lower overall aGVHD rate of 11.8%. Notably, all cases in this group were limited to grade I, whereas other studies have observed higher grades (II–IV or III–IV) of aGVHD.

Combining ATG with PTCy shows promise in improving GVHD outcomes, particularly by reducing moderate cGVHD. The absence of a statistically significant difference in aGVHD rates in both groups may be due to the limited sample size. Low-dose ATG targets early activated T-lymphocytes, while PTCy administered on day +3 eliminates rapidly proliferating T cells exposed to antigens and promotes the expansion of regulatory T cells. This complementary mechanism likely works synergistically, reducing GVHD risk through distinct but overlapping T-cell depletion pathways.^27–29

Relapse continues to be a significant challenge following haplo-HSCT. In this study, relapse rates at 1 and 2 years were comparable between groups. Specifically, the 1-year relapse incidence was 11.8% in the ATG/PTCy group and 6.3% in the PTCy group (p = 1.00), while 2-year rates were 17.6% and 12.5%, respectively (p = 1.00). Studies have demonstrated that adding ATG to standard GVHD prophylaxis does not increase relapse risk.^30,31 In this study, relapse was comparable between the low-dose ATG/PTCy group and the PTCy group (17.6% vs 12.5%; p = 1.00). Similarly, a meta-analysis by Luo et al.²⁶ found no significant increase in relapse risk with ATG/PTCy and suggested a potential trend toward lower relapse rates compared to PTCy alone. This consistency in relapse outcomes with low-dose ATG/PTCy may stem from optimized dosing strategies. Moreover, PTCy preserves CD8+ effector cells vital for the graft-versus-leukemia effect,^32,33 while ATG contributes by inducing apoptosis and complement-mediated lysis of leukemic cells, further supporting relapse prevention.³⁴

CMV DNAemia is a frequent complication in haplo-HSCT, with a cumulative incidence of 63.7%–66.1%, leading to considerable morbidity and mortality.^35,36 In this study, although not statistically significant, the low-dose ATG/PTCy group demonstrated a lower rate of CMV reactivation compared to the PTCy group (52.9% vs 81.3%). Consistent with this finding, Li et al.²⁴ observed a CMV reactivation rate of 51.4% in their evaluation of low-dose ATG/PTCy as a prophylaxis regimen. This lower rate of viral reactivation in low-dose ATG/PTCy regimen may be associated with the faster recovery of CD4+ T cells.

NRM was significantly lower in the low-dose ATG/PTCy group compared to the PTCy group in this study (17.6% vs 56.3%; p = 0.021). When analyzed over time, 1-year NRM was 11.8% in the ATG/PTCy group and 43.8% in the PTCy group (p = 0.057), while 2-year NRM rates reached statistical significance (17.6% vs 56.3%; p = 0.021). These results are consistent with findings from other research, including Yang et al.’s phase II trial, which reported an NRM rate of 9.4%.¹⁰ Similarly, Li et al.³⁷ and Wang et al.²⁸ documented NRM rates of 15.8%, 12.9%, and 6.0%, respectively, highlighting the lower mortality risk associated with the ATG and PTCy combination in haplo-HSCT. These results support the safety of this regimen, showing no adverse impact on mortality outcomes.

In terms of survival, OS and DFS were longer in the low-dose ATG/PTCy group compared to the PTCy group (70.6% vs 56.3%, and 64.7% vs 50% respectively). However, this difference was not statistically significant, likely due to the small sample size.

A meta-analysis by Luo et al.²⁶ found significantly higher OS rates for low-dose ATG/PTCy when compared to ATG alone (p = 0.02) or PTCy alone (p = 0.0006). Similarly, high 1-year OS rates of 80.0% and 74.9% were observed by Li et al.²⁴ and Xu et al.,²⁵ respectively, in patients receiving low-dose ATG/PTCy, aligning with the results of this study. In addition, Luo et al.²⁶ similarly reported significant DFS improvements with low-dose ATG/PTCy relative to ATG alone. The observed advantages in OS and DFS may result from the lower rates of cGVHD and NRM associated with the low-dose ATG/PTCy protocol. Further research is warranted to confirm these outcomes.

The primary limitations of this study include its limited sample size and retrospective nature, which may affect the generalizability of the findings. One limitation of this study is the absence of an a priori power analysis, as all eligible patients within the specified timeframe were included retrospectively, which may limit the generalizability and statistical power of some subgroup comparisons. However, the limited sample size warrants caution in interpreting these findings, and larger prospective trials are necessary to confirm these observations.

Conclusion

In conclusion, the combination of low-dose ATG and PTCy for GVHD prophylaxis in haplo-HSCT shows promise in significantly reducing the risk of moderate cGVHD and lowering NRM compared to PTCy alone. To our knowledge, this study represents the first direct comparison between low-dose ATG/PTCy and PTCy-only regimens. Larger, prospective trials are essential to confirm these findings and determine the optimal dosing strategies for this innovative GVHD prophylaxis approach.

Supplemental Material

sj-doc-1-tah-10.1177_20406207251353011 – Supplemental material for Low-dose anti-thymocyte globulin combined with low-dose post-transplant cyclophosphamide: a novel approach to prevent graft-versus-host disease in haploidentical stem cell transplantation

Supplemental material, sj-doc-1-tah-10.1177_20406207251353011 for Low-dose anti-thymocyte globulin combined with low-dose post-transplant cyclophosphamide: a novel approach to prevent graft-versus-host disease in haploidentical stem cell transplantation by Neslihan Mandaci Ṣanli and Ali Ünal in Therapeutic Advances in Hematology

Footnotes

Acknowledgements

None.

Declarations

ORCID iD

Neslihan Mandaci Ṣanli

Supplemental material

Supplemental material for this article is available online.

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