Virtual crossmatch–prodigal saviour of cardiac transplants in India

Abstract

Positive anti-human leukocyte antigen (HLA) antibodies are considered a contraindication for cardiac transplantation in India, due to its negative impact on cardiac allograft survival and increased chances of rejection post-transplant. Single antigen bead (SAB) assay helps to further characterize these antibodies. Our case portrays India’s first reported successful cardiac transplant utilizing a virtual crossmatch (VXM)-based approach in a patient with positive anti-HLA antibody screen and SAB assay. We propose that adopting such an approach, in select cases, in India is certainly feasible. This approach would lead to increasing the potential donor pool and mitigate the chances of post-transplant rejection. With increased demands, the SAB assay cost would reduce, further improving its cost-effectiveness.

Introduction

Anti-human leukocyte antigen (HLA) antibody screening is used to detect circulating anti-HLA antibodies in a transplant recipient’s serum. Pre-transplant anti-HLA antibodies can adversely impact cardiac allograft survival and rejection rates post-transplant [1]. A positive antibody screen also reduces the potential donor pool, thus increasing transplant wait time and wait-list mortality.

We present a cardiac transplant case with multiple pre-transplant antibodies, which was successfully managed utilizing “virtual” crossmatch (VXM) to optimize donor selection and pre-transplant de-sensitization to reduce the antibody burden.

Case report

A 62-year-old man with idiopathic restrictive cardiomyopathy was referred to our center with worsening New York Heart Association functional class IIIb dyspnea and multiple heart failure hospitalizations over 2 years. Echocardiogram demonstrated severe bi-ventricular dysfunction with restrictive physiology.

Right heart catheterization demonstrated severely reduced cardiac output (Fick 2.86 L/min) and acceptable pulmonary vascular resistance (3.1 WU). HLA antibody screening was positive for class 1 antibodies. In view of the positive anti-HLA antibody report, medical therapy was continued. The relatively small size of left ventricle and severe right ventricular dysfunction ruled out left ventricular assist device (LVAD) as a therapeutic option.

Patient declined clinically over next 6 months despite optimum medical therapy. Given the prior positive HLA antibody screen, a single antigen bead (SAB) assay was performed. The SAB assay (Immucor, USA) identified multiple class I anti-HLA antibodies (Mean Florescent Intensity {MFI} above 2000) and a single class II anti-HLA antibody (Table 1). Citing lack of other viable therapeutic modalities, we listed the patient for cardiac transplantation.

Table 1.

Single antigen bead (SAB) assay report (Luminex) with multiple un-acceptable class I and class II HLA antibodies in bold (MFI > 2000)

HLA class I antibodies	MFI value
HLA-A*33:01	6670
HLA-A*26:01	4908
HLA-A*66:02	3803
HLA-B*08:01	3362
HLA-A*33:03	2981
HLA-A*66:01	2218
HLA-A*69:01	2212
HLA-A*25:01	2181
HLA-A*68:02	1863
HLA-B*59:01	1532
HLA class II antibody	MFI value
HLA-DQB1*04:01	2189

Within 2 months of listing, we received a favorable ABO-compatible deceased donor offer. Donor HLA antigen profile testing (Table 2) along with complement-dependent cytotoxicity (CDC) and Luminex platform-based crossmatches were performed prospectively. Virtual crossmatch using recipient SAB result and the donor HLA profile was performed and it revealed absence of donor-specific antibodies (DSA) at the allelic level. Finally, with the negative results of CDC, Luminex, and virtual crossmatch, the donor was deemed acceptable.

Table 2.

Donor HLA profile without any un-acceptable antigens when crossmatched with antibodies in Table 1

A*68:01	B*51:01
B*07:05	C*14:02
C*07:01	DRB1*15:01
DRB1*04:04	DQB1*06:01
DQB1*03:02

Plasmapheresis was performed in the operating room, prior to the initiation of cardiopulmonary bypass. Intravenous rabbit anti-thymocyte globulin was used for induction immunosuppression. Transesophageal echocardiogram demonstrated normal bi-ventricular function. Post-operative plasmapheresis was not performed in view of negative DSA.

Patient was discharged home on standard maintenance immunosuppression comprising tacrolimus, prednisone, and mycophenolate mofetil. Endomyocardial biopsy performed at day 29 was negative for rejection. Repeat DSA testing was negative for de novo antibodies at 4 weeks, 3 months, 6 months, and 12 months post-transplant. Patient currently remains clinically stable, 13 months post-transplantation, with normal cardiac allograft function, without any episodes of cardiac rejection.

Discussion

Importance of anti-HLA antibodies for the survival of a transplanted kidney was first recognized by Patel et al. in 1969 [2]. This landmark study resulted in the routine use of CDC crossmatch in transplant patients to prevent hyperacute rejection and acute accelerated rejection. However, it is the least sensitive and specific of the current assays [3]. No single strategy for detecting pre-transplant antibodies is considered optimum, due to availability of multiple assays/platforms, with varying sensitivities and specificities and lack of large randomized controlled trials.

The International Society of Heart and Lung Transplantation (ISHLT) recommends (class IC) pre-transplant antibody screening in all prospective cardiac transplant candidates. If the antibody screening is positive, then further evaluation using SAB assay, solid-phase assays (flow cytometry or Luminex), and C1q assay is recommended [3]. However, the C1q assay is not readily available in India at this time.

According to the ISHLT guidelines, a VXM must be performed before a donor organ is allocated to a particular recipient [3, 4]. Currently in India, cardiac organ allocations occur on the basis of a single compatible ABO blood group crossmatch. The CDC, Luminex, and/or flow cytometry crossmatches are performed prior to the transplant. However, their results are available to the recipient team only when the cardiac transplant surgery is nearing completion or post-surgery.

Routine pre-transplant VXM requires close coordination and communication between the transplant team and histocompatibility team. It is ideal to perform flow cytometry or Luminex crossmatch along with the SAB assay. Often in India, VXM cannot be performed prior to the transplant due to time or infrastructure constraints. In such cases, we recommend performing HLA antibody screening, in addition to the CDC and Luminex crossmatch, with or without flow cytometry crossmatch, to rule out presence of DSA, which are detrimental to transplant outcomes. On the other hand, if a VXM is performed and is positive, then it is imperative to perform a prospective Luminex crossmatch and only proceed with transplant if the prospective crossmatch is negative.

The detection of anti-HLA antibodies is a crucial part of the evaluation and immune-monitoring of cardiac transplant recipients. The introduction of beads coated with HLA antigens has revolutionized the process of detection of these pre- and post-transplant antibodies. Luminex SAB assays are the most sensitive and specific of the bead assays [3]. The SAB assay results are expressed in terms of mean fluorescence intensity (MFI). In the absence of international standards, each transplant center has defined their own levels of accepted MFIs.

Immucor or One Lambda assay kits are commercially available and are widely used for performing the SAB assays. Studies comparing these assay kits showed similar results, especially in patients with moderate to high MFI [5, 6]. We acknowledge the known limitations of the SAB assays, such as the ability to detect antibodies that are reactive only to the antigens coated on the beads, without truly knowing the antigenic make-up of the Indian population pool. Further, in highly sensitized patients with high MFI values of more than 15,000 for several beads, it is necessary to dilute the sera for appropriate identification of antibodies. Thus, patients with low-avidity IgG and low-titer IgG can be missed in SAB assays in such instances.

Our patient demonstrated multiple pre-transplant anti-HLA antibodies. As explained above, cardiac transplant was the only remaining viable option. Meticulous donor selection utilizing VXM and planned pre-transplant de-sensitization strategies ensured a successful cardiac transplant.

Instead of an immunologically blind approach, we propose using a VXM-based approach in India, in this select group of patients, to help maximize our donor pool and mitigate rejection risk. Our case report is India’s first reported successful cardiac transplant case utilizing a VXM-based approach in a patient with significant pre-transplant antibodies with positive HLA antibody screening and SAB assay. With increased demands, the SAB assay costs would reduce further, improving its cost-effectiveness.

Funding

No funding was received to assist with the preparation of this manuscript.

Compliance with the ethical standards

Conflict of interest

The authors declare that they have no conflicts of interest.

Ethics committee approval

Approval from the institute ethics committee has been obtained. The authors declare that the clinical procedures involved were performed in accordance with the ethical standards as laid down in the 1964 Declaration of Helsinki and its later amendments.

Informed consent

Written, informed consent has been obtained from the patient.