Martin Gyger

Persistence of host Langerhans cells following allogeneic bone marrow transplantation: possible relationship with acute graft-versus-host disease

Langerhans cells (LC) are bone marrow‐derived dendritic antigen‐presenting cells found in the epidermis. In an effort to determine the origin (host versus donor) of LC at different intervals following bone marrow transplantation, we performed skin biopsies in 16 recipients of sex‐mismatched marrow. LC were identified using monoclonal antibody OKT6 in an indirect immunoperoxidase assay and their donor or host origin determined according to the presence or absence of Y body. The presence of Y‐positive (donor) LC could be demonstrated in all (6/6) skin biopsies of female recipients of male marrow tested between days 39 and 730 post‐transplant. Persistence of host LC in male recipients of female marrow was documented in all (6/6) recipients studied on day 39 and in two out of seven patients tested on day 120 post‐transplant. From day 365 onward, no residual host LC could be detected, suggesting that by this time all epidermal LC are donor‐derived. Our study demonstrates that host LC usually persist for 39 and up to 120 d following bone marrow transplantation. The relevance of this observation to the possible role of LC and other host dendritic antigen‐presenting cells in the graft‐versus‐host reaction is discussed.

Distinct patterns of minimal residual disease associated with graft-versus-host disease after allogeneic bone marrow transplantation for chronic myelogenous leukemia.

PURPOSE Allogeneic bone marrow transplantation (BMT) has been shown to provide effective therapy for chronic myelogenous leukemia (CML), but previous reports have also demonstrated the persistence of bcr-abl-positive cells for months to years after BMT in the majority of patients. To evaluate the biologic significance of persistent bcr-abl-positive cells, we examined the relationship between clinical parameters known to affect the risk of relapse and the ability to detect bcr-abl-positive cells post-BMT. PATIENTS AND METHODS We analyzed 480 samples from 92 patients at two transplant centers for the presence of bcr-abl-positive cells by polymerase chain reaction (PCR). Two different BMT preparative regimens and protocols for prevention of graft-versus-host disease (GVHD) were used. One center used cyclophosphamide plus total-body irradiation (CY/TBI) and T-cell-depleted marrow; the second center used busulfan plus cyclophosphamide (Bu/CY) and untreated marrow with cyclosporine and methotrexate (Csp/MTX) as GVHD prophylaxis. RESULTS We first determined the percent of patients at each center with > or = one PCR-positive (PCR+) result at defined intervals post-BMT. Between 0 and 6 months post-BMT, the majority of patients (80% to 83%) in both populations had PCR-detectable bcr-abl-positive cells. Between 6 and 24 months post-BMT, 80% to 88% of patients who received T-cell-depleted marrow remained PCR+, as compared with 26% to 30% of patients who received unmodified marrow. After 24 months post-BMT, the percentage of PCR+ patients was not significantly different in the two populations. This pattern of detection of bcr-abl-positive cells post-BMT followed the development of chronic GVHD in patients who received unmodified marrow. All patients were also divided into three groups based on post-BMT PCR results as follows: (1) persistent PCR+ (n = 29), (2) intermittent PCR-negative ([PCR-] n = 40), and (3) persistent PCR- (n = 23). These three groups were found to have a low, intermediate, and high probability of maintaining remission and disease-free survival, respectively (P = .0001). Intermittent or persistent PCR- results, which reflect levels of minimal residual disease < or = the limit of detection by PCR, were clearly associated with both acute (P = .004) and chronic (P = .000005) GVHD. Nevertheless, 44% of patients without GVHD also had intermittent or persistent PCR- assays. CONCLUSION The persistence of PCR-detectable bcr-abl-positive cells early post-BMT in more than 80% of patients suggests that neither BMT preparative regimen effectively eradicates CML cells in most patients. Subsequently, acute and/or chronic GVHD are associated with a decreased ability to detect residual bcr-abl-positive cells, which suggests that immunologic mechanisms mediated by donor cells are important for inducing long-term remissions after BMT. The demonstration that 44% of patients without GVHD had either low or undetectable levels of residual leukemia suggests the presence of mechanisms capable of suppression or eradication of CML independent of GVHD.

Immunodominant minor histocompatibility antigens expressed by mouse leukemic cells can serve as effective targets for T cell immunotherapy.

Numerous minor histocompatibility antigens (MiHAs) show tissue-specific expression and can induce vigorous T cell responses. They therefore represent attractive targets for leukemia immunotherapy mediated by adoptive transfer of T cells. The main objective of this work was to determine whether MiHAs expressed by normal hematopoietic cells were present on leukemic cells and whether they could trigger lysis by cytotoxic T lymphocytes (CTLs). CTL assays showed that mouse leukemic cells of both lymphoid and myeloid lineages were sensitive to CTLs targeted toward some but not all MiHAs. In four out of four strain combinations in which we primed CTLs against immunodominant MiHAs, effectors killed leukemic blasts, whereas no cytotoxicity was observed when CTLs were targeted toward four immunorecessive MiHAs. Testing of HPLC fractions obtained from normal and leukemic cells provided molecular evidence that leukemic blasts expressed only some of the MiHAs found on normal mouse hematopoietic cells. Decreased density of H-2 class I molecules at the surface of leukemic cells suggests that down-regulation of genes encoding either class I molecules or proteins involved in antigen processing played a role in the aberrant expression of MiHAs. In vivo resistance to the leukemic cells by various strains of mice correlated with in vitro CTL activity. These results show that leukemic cells express only some (immunodominant) MiHAs and suggest that this subset of MiHAs represent prime targets for adoptive immunotherapy.

Lymphoid interstitial pneumonia after allogeneic bone marrow transplantation. A possible manifestation of chronic graft‐versus‐host disease

Interstitial pneumonia (IP) is a frequent and serious complication of bone marrow transplantation with a median time of onset about 2 months posttransplant. Most cases result either from toxicity of radiation and chemotherapy or from infection with pathogens such as cytomegalovirus. Described are two patients with chronic graft‐versus‐host disease (GVHD) who presented with late‐onset IP 242 and 632 days posttransplant. Histologic examination of lung biopsy specimens disclosed a lymphoid interstitial pneumonia (LIP) in both cases. The major lymphocyte subset found in bronchoalveolar lavages and lung tissue was OKT8(+) and showed a positive dot staining for acid phosphatase. Contrary to peripheral blood mononuclear cells, most OKT8(+) lymphocytes in the lungs were OKT3(‐). Since acute GVHD lesions are mediated mainly by cytotoxic T‐lymphocytes, our data suggest that LIP in marrow‐grafted patients may be a manifestation of chronic GVHD. It should be distinguished from the more common types of IP encountered following bone marrow transplantation.

A case of pulmonary alveolar proteinosis complicating chronic myelogenous leukemia a peculiar pathologic aspect of busulfan lung

A 34‐year‐old woman with typical chronic myelogenous leukemia was treated with daily busulfan (total dose, 1600 mg approximately) from July 1978 to June 1981. In February 1981, she noticed a progressive deterioration of her clinical status, characterized by increasing dyspnea and productive cough. In July 1981, an open lung biopsy revealed pulmonary alveolar proteinosis. The patient died of progressive respiratory failure in August 1981. The association between chronic myelogenous leukemia and pulmonary alveolar proteinosis is briefly reviewed. The report discusses the possible etiologic role of busulfan therapy in the development of pulmonary alveolar proteinosis.

Immunobiology of allogeneic peripheral blood mononuclear cells mobilized with granulocyte-colony stimulating factor

The use of mobilized peripheral blood (PB) stem cells for autologous transplantation initially generated much enthusiasm because of enhanced engraftment in comparison to marrow stem cells and avoidance of general anesthesia for the donor. Its application to the allogeneic setting seemed inevitable. For obvious ethical reasons, allogeneic donors are mobilized with cytokines only, mainly granulocyte colony-stimulating factor (G-CSF). Results from preliminary studies suggest that in comparison to standard bone marrow transplants, outcomes such as engraftment, host-versus-graft reaction, graft-versus-host disease, graft-versus-leukemia and immunological reconstitution may be different. Surprisingly, G-CSF, previously recognized as a late acting lineage-specific factor for neutrophil production, not only disrupts homeostasis between stem cells and their microenvironment, but also induces significant quantitative and qualitative changes in the accessory cell compartment, affecting lymphocytes, monocytes, natural killer, dendritic, and stromal cells. Furthermore, mobilization of huge numbers of non-professional antigen presenting cells (CD34+ stem cells) amplifies the tolerizing potential of PB stem cell grafts. Thus, G-CSF mobilization provides PB transplants with different immunobiologic properties in comparison to standard bone marrow grafts. Whether these immunobiologic differences will lead to better transplant outcomes remains to be shown through much awaited results of large randomized clinical trials. Bone Marrow Transplantation (2000) 26, 1–16.

Bone marrow transplantation for myelodysplastic syndromes

Summary. Eight patients with myelodysplastic syndromes (MDS) were treated with bone marrow transplantation (BMT). Median age was 34.5 years and ranged between 3 and 45. FAB diagnosis was refractory anaemia (RA) in three, RA with excess of blasts (RAEB) in four and RAEB in transformation (RAEB‐t) in one case. Four patients were prepared with cyclophosphamide and total body irradiation whilst the other four received busulphan and cyclophosphamide. Engraftment was documented in seven of eight patients. Two patients died from complications related to the procedure. One had early veno‐occlusive disease of the liver whilst the other died 46 months after BMT from pulmonary fibrosis. One patient died from recurrent disease 11 months after BMT. Five patients are alive and in complete remission 9–35 months post‐transplantation. Four of these patients have a Karnofsky score ≤ 90%. These results suggest that BMT can induce prolonged disease‐free survival in patients under 50 years of age. If a compatible donor is available, marrow transplantation should be seriously considered in the treatment of MDS.

Chronic B-cell lymphocytosis

Persistent elevation of lymphocyte counts is usually associated with a malignant monoclonal lymphoproliferative disease. Over the last 8 years, amongst patients investigated in our center for undetermined persistent lymphocytosis, a diagnosis of malignant lymphoproliferation was excluded in 6 cases as studies of surface membrane immunoglobulin light chains showed that they presented a polyclonal expansion of their B‐lymphocyte pool. All patients were young‐to‐middle aged women presenting peculiar immunohematologic findings characterized by 1) persistent (2–7 yr) elevation of lymphocyte counts (4–14 × 109/l), 2) presence of characteristic binucleated B cells on peripheral blood smears, 3) a normal bone marrow histology, 4) a polyclonal increase of serum IgM with low‐to‐normal IgG and IgA levels. Histologic examination of the spleen in 2 patients and lymph nodes in 1 showed a benign follicular lymphoid hyperplasia. The evolution was benign in every case. We suggest that chronic polyclonal B‐cell lymphocytosis is a distinct clinicopathologic entity that should not be confused with malignant lymphoproliferative disorders.

Serum immunoglobulin levels following allogeneic bone marrow transplantation

SummaryIn order to study the posttransplant evolution of serum immunoglobulin levels, we measured serum IgG, IgA and IgM levels in 50 recipients of allogeneic bone marrow before transplantation and at different intervals thereafter (days 39, 120, 365 and 730). IgG and IgM levels were depressed for 1 year and IgA levels for 2 years posttransplant. Immunoglobulin deficiency was more severe and prolonged in patients with graft versus-host-disease. Hypogammaglobulinemia may contribute to the frequent infections observed in these patients, especially those with chronic graft-versus-host disease.

Therapy‐induced preleukaemia in patients treated for Hodgkin's lymphoma: clinical and therapeutic relevance of sequential chromosome banding studies

Between January 1978 and December 1982 successful sequential chromosome analyses were carried out on bone marrow cells of five patients previously treated for Hodgkins lymphoma (HL) presenting unexplained cytopenia or pancytopenia during follow‐up. All patients had concurrent morphological examination of bone marrow specimens showing signs of dysplasia and/or hypoplasia, without leukaemic infiltrate. Six other patients treated for HL who had normal haematological parameters served as controls. All the patients with unexplained cytopenias had clonal chromosome abnormalities; monosomy for chromosome No. 5 was the most frequent. No abnormalities were detected in the control group. Two patients have evolved to resistant leukaemia, one died of sepsis before leukaemic conversion while severely neutropenic, and two are in full marrow and cytogenetic recovery after aggressive anti‐leukaemic treatment in the pre‐leuk‐aemic phase. Our data suggest that cytogenetic studies may be of crucial value in detecting therapy‐induced preleukaemia (t‐PL) at an early stage of its evolution and in planning appropriate therapy before the establishment of overt leukaemia.