A successful haploidentical bone marrow transplantation method in rabbits: Perfusion method plus intra-bone marrow–bone marrow transplantation

Abstract

Graft versus host disease (GVHD), rejection, delayed immune reconstitution and infections have been significant hurdles to haploidentical BMT. In order to improve the outcome of the current haploidentical-related BMT, we performed a novel BMT method consisting of the perfusion method (PM) plus intra-bone marrow–bone marrow transplantation (IBM-BMT) in a rabbit model. The percentages of T cells in BMCs harvested by the PM and the conventional aspiration method (AM) were 6% and 14%, respectively (p < 0.01). Conversely, the CFU-C counts of BMCs in the PM group were significantly higher than those in the AM group. When the BMCs were transplanted into lethally irradiated offspring rabbits by IBM-BMT, hemopoietic recovery in the PM group was faster than in the AM group. The cumulative incidence of acute GVHD was 25% in the PM group versus 75% in the AM group (p < 0.05). In addition, the survival rate was 75% in the PM group versus 33% in the AM group (p < 0.05). Thus, the new method is able to provide rapid hemopoiesis, reduce the cumulative incidence of acute GVHD, and achieve a higher survival rate. This novel strategy paves the way for new dimensions in haploidentical BMT.

Introduction

Allogeneic hemopoietic stem cell transplantation (HSCT) is one of the definitive therapies for advanced hemopoietic malignancies. The success of allogeneic HSCT depends entirely on the presence of a suitable donor with fully matched or acceptably mismatched human leukocyte antigen (HLA). In the absence of an HLA-matched sibling, patients need to have an HLA-matched-unrelated donor's BMCs or cord blood cells. However, the timing of transplantation is often influenced by the time-consuming procedure of a marrow donor program. In the case of cord blood cells, graft failure is sometimes induced by the insufficient concentration of the hemopoietic stem cells (HSCs). In the case of BMCs, to prevent acute GVHD, T-cell-depleted or CD34⁺-selected HSCT has been extensively attempted during the past decade. However, transplantation frequently failed due to infection and leukemia relapse [1], [2], [3], [4], [5], [6], [7], [8], [9], [10], [11]. Recently, we developed a novel BMT method, which combines an advanced BMC-harvesting method (PM) with IBM-BMT [12]. Compared with the conventional AM, the PM, which has been performed on the long bones (humerus, femur and tibia) and also the iliac bones of cynomolgus monkeys, allows BMCs to be collected with minimal contamination with peripheral T cells [13], [14]. As a consequence, PM can significantly reduce the risk of GVHD. In the case of IBM-BMT, donor BMCs are directly injected into the recipient bone marrow cavity. Therefore, injected allogeneic donor cells are able to interact efficiently with donor-derived stromal cells, including mesenchymal stem cells (MSCs), which cause the proliferation, differentiation, and even maintenance of HSCs [15], [16]. In the case of conventional intravenous BMT (IV-BMT), the majority of the BMCs (both HSCs and MSCs) are trapped in the lung [17], [18], [19] and liver [20], where they are killed by the radio-resistant host cells. Compared with IV-BMT, this novel method generates an earlier engraftment of hematolymphoid cells of donor-origin [21]. Thus, we succeeded in treating the intractable autoimmune diseases in chimeric-resistant MRL/lpr mice by IBM-BMT. Following these results, in order to diminish the hurdles to allogeneic BMT, PM + IBM-BMT was performed on haploidentical-related rabbits. Due to their higher radiosensitivity [22], [23] and their resistance to inbreeding [24], rabbits are well-recognized as being difficult animals on which to perform allogeneic BMT. However, the genetic heterogeneity of a rabbit population makes the rabbit much more akin to human beings in terms of their biological individuality than the pure-bred homogeneous strains of mice usually used in these studies. Thus, we used the rabbit system to gauge the effectiveness of this novel BMT method. In this report, we demonstrate that PM + IBM-BMT is able to allow a wide spectrum of donor-matching, prevent the development of GVHD, improve the survival rate, and generate a persistent and stable chimera for at least 2 years. Accordingly, this novel approach might deliver the expected innovative therapies for the various otherwise intractable diseases in humans, including autoimmune diseases, genetic disorders, and leukemia.