A Pilot Randomized Controlled Trial of De Novo Belatacept-Based Immunosuppression after Lung Transplantation

Abstract

Background:

Chronic lung allograft dysfunction (CLAD) is the leading cause of death beyond the first year after lung transplantation. The development of donor-specific antibodies (DSA) is a recognized risk factor for CLAD. Based on experience in kidney transplantation, we hypothesized that Belatacept, a selective T-cell co-stimulatory blocker, would reduce the incidence of DSA after lung transplantation which may ameliorate the risk of CLAD.

Methods:

We conducted a pilot randomized-controlled trial (RCT) at 2 sites to assess the feasibility and inform the design of a large-scale RCT. All subjects were treated with rabbit anti-thymocyte globulin for induction immunosuppression. Subjects in the Control arm were treated with Tacrolimus, Mycophenolate Mofetil (MMF), and Prednisone, and subjects in the Belatacept arm were treated with Tacrolimus, Belatacept, and Prednisone through day 89 after transplant then converted to Belatacept, MMF, and Prednisone for the remainder of year 1.

Results:

After randomizing 27 subjects, 3 in the Belatacept arm died compared to none in the Control arm. As a result, we stopped enrollment and treatment with Belatacept, and all participants were treated with standard of care immunosuppression. Overall, 6 participants in the Belatacept arm died compared to none in the Control arm (log rank p = 0.008). We did not observe any differences in the incidence of DSA, acute cellular rejection, antibody-mediated rejection, CLAD, or infections between the 2 groups.

Conclusions:

We conclude that the investigational regimen used in this pilot RCT is associated with increased mortality after lung transplantation.

INTRODUCTION

Lung transplantation improves survival and quality of life for patients with advanced lung disease.^1–3 However, long-term outcomes remain disappointing, and the median survival after lung transplantation is 6.7 years.⁴ The leading cause of death beyond the first year after lung transplantation is chronic lung allograft dysfunction (CLAD)⁴. The development of donor-specific antibodies (DSA) to mismatched human leukocyte antigens (HLA) is widely recognized as an independent risk factor for CLAD and death after lung transplantation,^5–7 and DSA cause antibody-mediated rejection (AMR) which commonly leads to CLAD and allograft failure.^8–12 Moreover, the development of DSA is common after lung transplantation. In a prospective multicenter observational study, 36% of lung recipients developed DSA within 120 days of transplantation.¹³ In other studies, the incidence of DSA within the first year after transplantation is approximately 50%.^6,7 These findings illustrate that current immunosuppressive regimens do not sufficiently prevent the development of DSA or their deleterious effects on the lung allograft.

Belatacept, a CTL4-Ig fusion protein, is a novel immunosuppressant that binds CD80 and CD86 thereby blocking CD28 co-stimulatory signals.^14–17 Belatacept is approved by the Food and Drug Administration (FDA) for the prevention of rejection after kidney transplantation. In a multicenter randomized-controlled trial (RCT), kidney transplant recipients treated with Belatacept were less likely to develop DSA and had better survival than those treated with Cyclosporine, but there was no significant difference in the incidence of serious adverse events (SAE) or serious infections between the 2 groups.¹⁷ Based on these data, we hypothesized that Belatacept would prevent the development of DSA and CLAD after lung transplantation and conducted a pilot RCT to assess the feasibility and inform the design of a phase 3 RCT that would examine the efficacy and safety of Belatacept in lung transplantation.¹⁸ Enrollment, randomization, and treatment in the pilot RCT were stopped after 3 subjects randomized to Belatacept died compared to none of the Control subjects.¹⁸ All subjects in the Belatacept arm were converted to standard of care immunosuppression, and follow-up was continued for 14 months after randomization. We previously reported the increased mortality to alert clinicians and investigators; here, we report final results of the study after completion of follow-up.

MATERIALS AND METHODS

Study design and medical regimen

We previously detailed the study design and medical regimen.¹⁴ Briefly, we performed a pilot 2-center phase 2 RCT to assess the feasibility of conducting a large-scale RCT that would examine the efficacy and safety of Belatacept after lung transplantation. The primary endpoint of the pilot study was the feasibility metric of randomizing 80% of eligible patients within 4 hours of completion of transplantation. The study protocol and additional supporting information may be found in the Supplemental Digital Content. We enrolled subjects after listing for transplantation and randomized eligible participants after transplantation using a computer-generated block randomization method with a 1:1 ratio to either the Belatacept arm or the Control arm. All subjects were treated with rabbit anti-thymocyte globulin (ATG) 1 mg/kg on days 0, 1, and 2 for induction immunosuppression. Subjects in the Control arm were treated with Tacrolimus, Mycophenolate Mofetil (MMF), and prednisone through the study period. Subjects in the Belatacept arm were treated with Belatacept, Tacrolimus, and prednisone starting on day 0 through day 89. On day 90, Tacrolimus was replaced by MMF and Belatacept and prednisone were continued through day 365. In both arms, Tacrolimus was initiated enterally or sublingually with the first 48 hours after transplantation and dosed to target a trough blood level of 8–15 ng/mL, and MMF was dosed at 1 g twice daily. In the Belatacept arm, the first dose of Belatacept was given after transplantation 12 hours after the dose of ATG to avoid thrombotic complications. Belatacept was dosed at 10 mg/kg on days 0, 7, 14, 28, 56, and 84, then at 5 mg/kg on days 112, 140, 168, 196, 224, 252, 280, 308, 336, and 364. All patients were treated with methylprednisolone 500 mg intravenously before reperfusion of the allograft during the transplant surgery. After transplant, participants were treated with methylprednisolone 0.5 mg/kg intravenously twice daily for 6 doses, then prednisone 0.5 mg/kg daily through day 14, then 0.2 mg/kg daily through day 30, then 0.1 mg/kg daily through day 180, then 5 mg daily through day 365. All participants who were at risk for cytomegalovirus (CMV) infection (i.e., CMV seronegative recipients of seropositive donors or CMV seropositive recipients) were treated with Valganciclovir prophylaxis through day 365. All participants received appropriate prophylaxis against Pneumocystis jirovecii. Antifungal prophylaxis was initiated based on culture results from bronchoscopy specimens.

HLA antibody testing was conducted at the study core lab at the Baylor University Medical Center Transplant Immunology Laboratory, and the HLA investigator and technicians were blinded to participants’ study arm assignment. Details regarding HLA antibody testing are provided in the study protocol in the Supplemental Digital Content. All lung biopsies were interpreted at the study core pathology lab at Washington University by a single pathologist who was also blinded to participants’ study arm assignment.

Once enrollment and treatment with Belatacept were stopped, the primary endpoint of the study was changed to survival. Other clinical outcomes were assessed as secondary endpoints (Table S1 in the Supplemental Digital Content). Study participants randomized to Belatacept who were within the first 89 days of transplant when treatment was stopped were started on MMF 1 g twice daily, whereas those beyond day 90 were started on Tacrolimus to replace Belatacept. All participants in the Belatacept arm were receiving Belatacept every 28 days when the study status changed. Thus, Tacrolimus or MMF was started on day 21 ± 3 of the Belatacept treatment cycle to replace Belatacept and allow Tacrolimus or MMF to reach therapeutic levels as Belatacept’s effects and levels wane. Participants in the Control arm were continued on Tacrolimus, MMF, and prednisone. In the event of gastrointestinal toxicity, Azathioprine 2 mg/kg daily or enteric coated MMF 720 mg twice daily were substituted for MMF in both arms.

Safety monitoring and reporting and study oversight are detailed in the study protocol in the Supplemental Digital Content. Belatacept was used under an FDA Investigational New Drug (IND) application (138662). The sites’ Institutional Review Boards approved the study protocol and subsequent modifications, and the study was registered on ClinicalTrials.gov (NCT03388008).

Cellular immune phenotyping

To explore potential differences between the 2 study arms on cellular immune responses, we analyzed the frequency of NK cells, CD4⁺ and CD8⁺ T cell naïve and memory cell populations isolated from peripheral blood mononuclear cells (PBMCs) at serial timepoints. We analyzed memory T cell abundance given that memory T cell development initially requires CD28 engagement on naïve T cells. CD4⁺ and CD8⁺ T cells were stained with CD45RA and CCR7 to delineate naïve (CD45RA⁺ CCR7⁺), central memory (T_CM, CD45RA⁻ CCR7⁺), effector memory (T_EM, CD45RA⁻ CCR7⁻), and effector memory cells re-expressing CD45RA (T_EMRA, CD45RA⁺ CCR7⁻) subsets. NK cells were identified on a CD3⁻ CD8⁻ CD4⁻ CD56⁺ gate. The abundance of each lymphocyte subset was analyzed by flow cytometric analysis using a FACScan cytometer (Beckton Dickinson, Franklin Lakes, NJ USA. All antibodies were purchased from Biolegend (San Diego, CA USA).

Statistical analyses

We compared baseline characteristics between the 2 groups using t-tests or Wilcoxon-Rank sum tests if the data were not normally distributed and chi-square tests. We used the Kaplan-Meier method to report freedom DSA, ACR, CLAD, and survival and compared the outcomes between the 2 groups using the log rank test. We used univariate Cox proportional hazards models to explore baseline characteristics that may be associated with mortality. We conducted all analyses according to the intention to treat principle and used SPSS and Prism. We considered p < 0.05 statistically significant.

RESULTS

Between December 1, 2019, and May 30, 2021, we enrolled 49 subjects, and 27 were randomized: 13 were randomized to Belatacept and 14 were randomized to Control (Figure 1). All subjects who were eligible for randomization were randomized within 4 hours of completion of transplantation. Table 1 illustrates randomized participants’ baseline characteristics including metrics of disease severity, operative variables, and donor characteristics. Subjects randomized to Belatacept had significantly shorter 6-minute walk distances before transplantation, but there were no other significant differences between the 2 groups. Interstitial lung disease (ILD) was the leading indication for transplantation in both groups, and the majority of participants underwent bilateral lung transplantation. Additional detailed results are provided in the Supplemental Digital Content.

Figure 1.

Study consort diagram.

Table 1.

Baseline characteristics of randomized participants.

	Belatacept N = 13	Control N = 14	p value
Recipient age, mean ± SD, median, IQR	57.8 ± 7.5, 58, 13	59.5 ± 11.8, 62.5, 5	0.655
Recipient sex			0.785
Female, n (%)	4 (31%)	5 (36%)
Male, n (%)	9 (69%)	9 (64%)
Recipient race			0.586
Black, n (%)	1 (8%)	2 (14%)
White, n (%)	12 (92%)	12 (86%)
Recipient ethnicity			0.957
Hispanic, n (%)	1 (8%)	1 (7%)
Not Hispanic, n (%)	12 (92%)	13 (93%)
Diagnosis leading to transplantation			0.202
Interstitial lung disease, n (%)	6 (46%)	8 (57%)
Chronic obstructive pulmonary disease, n (%)	4 (31%)	0
Pulmonary arterial hypertension, n (%)	0	2 (14%)
Bronchiectasis, n (%)	1 (8%)	1 (7%)
Sarcoidosis, n (%)	0	1 (7%)
Other, n (%)	2 (15%)	2 (14%)
Operation			0.315
Single lung transplant, n (%)	1 (8%)	3 (21%)
Bilateral lung transplant, n (%)	12 (92%)	11 (79%)
Cardiopulmonary bypass, n (%)	0 (0%)	2 (14%)	0.157
Intra-operative extracorporeal membrane oxygenation support, n (%)	5 (38%)	6 (43%)	0.816
Cytomegalovirus serostatus			0.594
Recipient seronegative/Donor seronegative, n (%)	1 (8%)	3 (21%)
Recipient seronegative/Donor seropositive, n (%)	5 (38%)	5 (36%)
Recipient seropositive/Donor seropositive or seronegative, n (%)	7 (54%)	6 (43%)
Lung allocation score at transplant, mean ± SD, median, IQR	46.74 ± 17.23, 39.72, 19.43	38.69 ± 4.70, 37.15, 5.91	0.257
Hospitalized immediately before transplant, n (%)	4 (31%)	2 (14%)	0.303
Supplemental oxygen at rest before transplant (liters per minute), mean ± SD, median, IQR	7.6 ± 12.5, 2, 13	1.1 ± 1.4, 0, 2	0.209
Supplemental oxygen during exertion before transplant (liters per minute), mean ± SD, median, IQR	15.2 ± 16.3 8, 17	12.0 ± 9.0 10, 14	0.695
6-minute walk distance before transplant (feet), mean ± SD, median, IQR	612 ± 305, 710, 406	987 ± 376, 967, 344	0.005
Body mass index (kg/m2) before transplant, mean ± SD, median, IQR	25.06 ± 3.44, 25.15, 6.3	28.44 ± 2.68, 28.21, 3.4	0.370
Creatinine (mg/dL) before transplant, mean ± SD,  median, IQR	0.86 ± 0.21, 0.88, 0.39	0.92 ± 0.31, 0.85, 0.36	0.561
Mean pulmonary artery pressure (mm Hg) before transplant, mean ± SD	28 ± 11, 26.5, 16	34 ± 13, 29, 25	0.695
Cardiac index (L/min/m2) before transplant, mean ± SD, median, IQR	2.69 ± 0.46, 2.57, 0.73	2.69 ± 0.81, 2.45, 1.17	0.996
Donor age, mean ± SD, median, IQR	39.89 ± 15.15, 36, 25.21	32.20 ± 14.47, 30.5, 25.5	0.190
Donor sex			0.173
Female, n (%)	6 (46%)	3 (21%)
Male, n (%)	7 (54%)	11 (79%)
Donor smoking > 20 pack-years, n (%)	1 (8%)	1 (7%)	0.957
Final donor PaO2:FiO2 ratio before transplant, mean ± SD, median, IQR	419 ± 69, 414, 120	421 ± 63, 413, 92	0.959
Primary graft dysfunction grade at T0, n (%)			0.513
Grade 0	6 (46%)	10 (71%)
Grade 1	3 (23%)	1 (7%)
Grade 2	2 (15%)	1 (7%)
Grade 3	2 (15%)	2 (14%)

There was no significant difference in freedom from DSA with a Mean Fluorescence Intensity (MFI) threshold ≥ 2000 or 4000 between the 2 groups (Figure 2A and Figure 2C, respectively). Likewise, there was no significant difference in the distribution of DSA class using either MFI threshold between the 2 groups (Figure 2B and Figure 2D). Finally, there was no significant difference in freedom from complement-activating (C1q-positive) DSA or the distribution of C1q-positive DSA class between the 2 groups (Figure 2E and Figure 2F). No new DSA were identified in the Belatacept arm after participants were converted to standard of care immunosuppression. There was no significant difference in freedom from acute cellular rejection (ACR) grade A1 between the 2 groups (Figure 3A). Similarly, there was no significant difference in freedom from lymphocytic bronchiolitis (LB) grade B1R between the 2 groups (Figure 3B). Two participants in the Belatacept arm had an episode of ACR grade A1 after conversion to standard of care immunosuppression. Follow-up biopsies in both cases showed resolution of ACR, and neither participant developed CLAD during the study period. There were no cases of LB in the Belatacept arm after conversion to standard of care immunosuppression.

Figure 2.

The development of donor-specific antibodies (DSA). There was no significant difference in freedom from the development of DSA between the 2 groups.

Figure 3.

Acute cellular rejection (ACR) and lymphocytic bronchiolitis (LB). There was no significant difference in freedom from ACR or LB between the 2 groups.

One participant in each arm developed probable AMR according to the International Society for Heart and Lung Transplantation (ISHLT) criteria.¹⁹ The participant in the Belatacept arm developed CLAD after the diagnosis of AMR, but the participant in the Control arm recovered to their previous baseline. Both participants who developed AMR were alive at the end of study follow-up. There was no significant difference in freedom from CLAD between the 2 study arms (Figure 4A). Two participants in each arm developed CLAD. Of the 2 participants in the Belatacept arm who developed CLAD, 1 developed Restrictive Allograft Syndrome (RAS) 6 weeks after a respiratory infection with rhinovirus/enterovirus that was complicated by acute hypoxemic respiratory failure, and 1 developed CLAD with mixed features of Bronchiolitis Obliterans Syndrome (BOS) and RAS after the diagnosis of AMR. Both participants in the Control arm who developed CLAD developed BOS. One participant who developed RAS died of progressive CLAD whereas the other participants were alive at the end of study follow-up.

Figure 4.

A. Freedom from chronic lung allograft dysfunction (CLAD); there was no significant difference in freedom from CLAD between the 2 groups. B. Patient survival; there was a significantly worse survival in the Belatacept arm compared to the Control arm (log rank p = 0.008).

There was no significant difference in the incidence of infection between the 2 arms (Table 2). Three participants in each arm developed Coronavirus Disease 2019 (COVID-19) infection (Table 2). Two participants in each arm developed COVID-19 infection before the availability of vaccines, and 1 in each arm had been vaccinated but developed breakthrough infection. Two participants in the Belatacept arm died due to COVID-19 infection; 1 developed infection before the availability of vaccines, and 1 developed infection after receiving 3 doses of the BNT162b2 vaccine. Both had prolonged, refractory hypoxemic respiratory failure that led to death. One of these participants developed COVID-19 and hypoxemic respiratory failure that required hospitalization during study follow-up. She remained hospitalized with persistent respiratory failure and died after completing study follow-up. None of the participants in the Control arm died due to COVID-19 although 1 participant had 2 discrete COVID-19 infections. There were 5 cytomegalovirus (CMV) seronegative recipients of seropositive donors in each group (Table 1). Among these, 2 in the Belatacept arm and 1 in the Control arm had at least 1 breakthrough CMV infection; 1 of the Belatacept participants had protracted viremia with ganciclovir resistant CMV, but ultimately cleared the infection. The other CMV infections were uncomplicated. There were no significant differences in the incidences of chronic kidney disease stage 3, diabetes mellitus or hypercholesterolemia requiring medical therapy between the 2 groups (Table 3). As previously reported, 1 participant in the Belatacept arm developed Epstein-Barr Virus (EBV)-associated Post-Transplant Lymphoproliferative Disease (PTLD) that was fatal.¹⁸ This recipient was EBV seropositive before transplantation.

Table 2.

Infections

	Belatacept N = 13		Control N = 14		p valuea
	N affected	N events	N affected	N events
Any infection	11	35	12	50	0.936
Cytomegalovirus infection	6	9	4	6	0.345
CARVb infection	5	5	7	11	0.547
COVID-19c	3	3	3	4	0.918
Bacterial respiratory tract infection	7	10	10	14	0.345
Bacterial urinary tract infection	1	1	0	0	0.290
Fungal respiratory tract infection	3	4	7	12	0.148
Surgical site infection	1	1	1	1	0.957
HSVd respiratory or mucocutaneous infection	2	2	2	2	0.936

^aThe number of affected individuals is compared between the 2 groups.

^bCARV: community-acquired respiratory virus.

^cCOVID-19: Coronavirus Diseases 2019.

^dHSV: Herpes Simplex Virus.

Table 3.

Other complications of immunosuppression.

	Belatacept N = 13	Control N = 14	p value
Chronic kidney disease stage 3, n (%)	4 (31%)	3 (21%)	0.580
Diabetes mellitus requiring medical therapy, n (%)	7 (54%)	7 (50%)	0.842
Hypertension requiring medical therapy, n (%)	2 (15%)	7 (50%)	0.057
Hypercholesterolemia requiring medical therapy, n (%)	2 (15%)	2 (14%)	0.936
Malignancy, n (%)	1 (8%)	0 (0%)	0.290

Eleven participants in the Belatacept arm and 13 in the Control arm experienced at least 1 serious adverse event (SAE) during follow-up (p = 0.496). The most common SAE was acute hypoxemic respiratory failure (Table 4). Six participants in the Belatacept arm and 6 in the Control arm experienced at least 1 episode of acute hypoxemic respiratory failure (Table 4). One participant in the Belatacept arm experienced 6 episodes of respiratory failure. Common causes of acute hypoxemic respiratory failure in both arms included respiratory tract infection, early postoperative respiratory failure, and airway complications. Other SAE are listed in Table 4.

Table 4.

Serious adverse events.

	Belatacept N = 13		Control N = 14
	N affected	N events	N affected	N events
Acute hypoxemic respiratory failure	6	13	6	6
COVID-19 infection	3	3	3	4
Cytomegalovirus infection	2	3	1	1
Pneumonia or respiratory tract infection	1	2	2	2
Surgical site infection	1	1	1	1
Hemoptysis	1	1	0	0
Hemothorax	2	2	0	0
Atrial fibrillation	1	1	2	2
Dysphagia	0	0	1	1
Nausea/vomiting	2	2	0	0
Acute rejection	0	0	2	2
Polytrauma and fractures	1	1	0	0
Post-transplant lymphoproliferative disease	1	1	0	0
Headache	1	1	0	0
Syncope	0	0	1	1
Acute kidney injury	0	0	2	3
Limb ischemia	0	0	1	1
Venous thromboembolism	2	3	1	1
Sternal dehiscence	1	1	1	1
Cardiac arrest	1	1	0	0
Acute cholecystitis	0	0	1	1
Altered mental status	0	0	1	1
Feeding tube malfunction	0	0	1	2

Five participants in the Belatacept arm died during the study period, and a sixth participant died after completing study follow-up but had been hospitalized with prolonged respiratory failure after COVID-19 infection that occurred during the study period. In contrast, none of the participants in the Control arm died (Figure 4B, log rank p = 0.008). Causes of death included COVID-19 infection in 2 participants, CLAD in 1 participant, PTLD in 1 participant, hemothorax in 1 participant, and a suspected pulmonary embolism in 1 participant. None of the baseline recipient, donor, or operative characteristics was associated with mortality (Table S2).

We analyzed levels of NK, CD4+ and CD8+ T cell naïve and memory populations isolated from serial samples of PBMCs from 13 subjects (Control, n = 7 and Belatacept, n = 6). Lymphocyte levels were plotted as a function of time and interpolated by simple linear regression. Notably, there was a nonzero increase in NK cell abundance in the Belatacept arm (p = 0.003), which was absent in the control arm (p = 0.2) (Figure 5). However, we did not observe any differences in the frequency of naïve or central memory (T_CM), effector memory (T_EM) or effector memory re-expressing CD45RA (T_EMRA) cells between the 2 study arms at the different timepoints (Figure 5).

Figure 5.

Analyses of levels of NK, CD4+ and CD8+ naïve and memory T cell populations from peripheral blood mononuclear cells of 13 participants. There was a nonzero increase in NK cell abundance in the Belatacept arm which was absent in the control arm, but there were no significant differences in the frequency of CD4+ and CD8+ T cell naïve, central memory (T_CM), effector memory (T_EM), and effector memory cells expressing CD45RA (T_EMRA) between the 2 arms.

DISCUSSION

We designed this pilot study to assess the feasibility of conducting a phase 3 RCT examining the efficacy of Belatacept-based immunosuppression after lung transplantation. Although the study results demonstrate the feasibility of randomizing lung transplant recipients in the immediate postoperative period, we observed increased mortality with the investigational regimen which prompted termination of enrollment, randomization, and treatment. Unfortunately, 3 additional participants randomized to the investigational regimen died after conversion to standard of care immunosuppression for a total of 6 deaths in the Belatacept arm compared to none in the Control arm. As in our original Brief Communication, we conclude that the investigational regimen used in this study consisting of induction immunosuppression with rabbit ATG and this Belatacept dosing regimen is associated with increased mortality after lung transplantation.¹⁸

Although there were different causes of death, it is notable that 4 of the 6 deaths were associated with viruses; 2 deaths were due to COVID-19 infection, 1 was due to progressive RAS 6 weeks after rhinovirus/enterovirus infection, and 1 was due to EBV-associated PTLD in an EBV seropositive recipient. Some studies in kidney transplantation have linked Belatacept to an increased risk of CMV infection and disease particularly among CMV seronegative recipients of seropositive donors.^20–22 Thus, it is interesting that we observed increases in NK cell levels in Belatacept-treated participants which is possibly suggestive of ongoing sub-clinical viral infection. It is the prevailing view that Belatacept’s inhibition of the CD28-B7 costimulatory pathway is required for the development of primary T-cell responses but is less important for memory responses.^{22, 23} Indeed, Belatacept is contraindicated in EBV seronegative patients because of the increased risk of EBV-associated PTLD. Although we did not observe a difference in the number of CMV infections between the 2 study groups, even among CMV seronegative recipients of seropositive donors, the risk may have been ameliorated by prophylactic valganciclovir in all recipients at risk for CMV during the study period. In addition, the sample size of CMV seronegative recipients of seropositive donors was small in each study arm (n = 5). Nonetheless, Belatacept’s effect on naïve T-cell responses would not explain PTLD in our patient because they were EBV seropositive before transplantation. However, Belatacept’s inhibition of naïve T-cell responses may explain the observed 2 deaths related to COVID-19 and the death due to progressive RAS after rhinovirus/enterovirus infection in the Belatacept arm. Of the 6 total study participants who developed COVID-19, only 2 were vaccinated because most infections occurred before the availability of vaccines. Multiple reports have documented poor humoral and cellular responses to COVID-19 and influenza vaccination among kidney transplant recipients treated with Belatacept.^24–28 However, previous reports have not shown increased mortality due to COVID-19 among kidney transplant recipients treated with Belatacept.^29–30 Moreover, in a multi-center RCT of hospitalized nontransplant patients with moderate to severe COVID-19, a single dose of Abatacept, another CTL4-Ig that prevents CD28 binding to CD80/CD86, was associated with significantly better 28-day survival than placebo suggesting that co-stimulatory blockade may have favorable immune modulating effects in the setting of COVID-19 infection.³¹ However, chronic co-stimulatory blockade may result in different outcomes, and lung transplant recipients have a higher risk of death than nontransplant hospitalized patients with COVID-19.

Importantly, we did not detect any potential benefits attributable to treatment with Belatacept although the study was not powered to assess efficacy. There was no difference in the incidence of DSA between the 2 study groups. As in other studies, 14 of the 27 total subjects in this study developed DSA with an MFI threshold ≥ 2000.^6–7 However, it is noteworthy that all participants in this study were treated with rabbit ATG for induction immunosuppression suggesting that cytolytic induction immunosuppression may not impact the risk of DSA development after lung transplantation. We did not observe a difference in the incidence of AMR, ACR, LB, or CLAD between the 2 study groups. Although there were no clear differences in infections or SAE between the 2 groups, it is possible that our data underestimate the true incidence of these events because of the shorter duration of follow-up among those who died in the Belatacept arm. Finally, despite increases in NK cell levels, we did not observe any significant changes in the relative proportions of naïve and memory T cells in subjects in the Belatacept arm compared to those in the Control arm as function of time after transplant. Although these observations suggest that Belatacept treatment did not alter naïve or memory T cell maintenance further phenotypic and functional assays would be required to determine if there are pronounced deficits in T cell activation or memory cell development.

This study has several important limitations including the small sample size. Although our planned sample size was 40 total subjects, we halted enrollment, randomization, and treatment because of excess mortality in the investigational arm. Furthermore, the study was not powered to assess efficacy, and small but important differences in clinical outcomes other than survival may not have been detected. Such differences may explain the significantly worse survival in the Belatacept arm. However, small pilot studies that assess feasibility can inform the design of larger clinical trials that examine efficacy. Indeed, although this study did not show a clinical benefit, it prevented the initiation of a larger RCT that would have exposed more participants to this investigational regimen. This underscores the role of early-phase clinical trials of novel therapeutics even though these are not designed to assess efficacy and may not change clinical practice. Another important limitation is that our conclusion is specific to this investigational regimen, and other regimens using different Belatacept dosing or in combination with different immunosuppressants may have different results. However, we suspect that another clinical trial using Belatacept in lung transplantation may not be practical. Additional safety and compelling efficacy data, perhaps in a nonhuman primate model, would be necessary to justify another clinical trial of Belatacept in lung transplantation. Although some case series have reported a potential role for Belatacept in lung transplant recipients who are intolerant of calcineurin inhibitors, others have observed severe toxicity that led to treatment discontinuation.^32,33 In combination with our results, this suggests that there is an inherent risk to the use of Belatacept in lung transplantation although predictors of poor outcomes are unknown.

We conclude that the investigational immunosuppressive regimen used in this pilot RCT is associated with increased mortality after lung transplantation. These findings emphasize the need to examine novel immunosuppressive treatments in lung transplantation in the context of carefully designed and monitored clinical trials rather than incorporating these into clinical practice based on findings in other solid organ transplant settings.

ABBREVIATIONS PAGE

ACR

acute cellular rejection

AMR

antibody-mediated rejection

ATG

anti-thymocyte globulin

BOS

bronchiolitis obliterans syndrome

CLAD

chronic lung allograft dysfunction

DSA

donor-specific antibody

FDA

Food and Drug Administration

HLA

human leukocyte antigen

LB

lymphocytic bronchiolitis

MMF

mycophenolate mofetil

PBMCs

peripheral blood mononuclear cells

RAS

restrictive allograft syndrome

RCT

randomized-controlled trial

SAE

serious adverse event