Detection of minimal residual disease (MRD) after bone marrow transplantation (BMT) by multi-parameter flow cytometry (MPFC)

Multi-parameter flow cytometry (MPFC) was used to detect minimal residual disease (MRD) following bone marrow transplantation (BMT) in 21 patients. Bone marrow (BM) was analyzed pre-transplant and 3–4 months post-BMT while the patients were in clinical and morphological remission. MRD was detected by identifying cells with aberrant antigen expression and/or leukemia-associated phenotype (LAP) using MPFC. Prior to BMT, 8 out of 21 patients exhibited normal antigen expression based on normal BM samples while 13 BM aspirates had abnormal MPFC. Pre-BMT MPFC was abnormal in all 10 patients who were not in complete remission (CR) (>5% blasts in BM) as well as 3 patients acute lymphoblastic leukemia (ALL) who were in CR. In BM from ALL patients, an abnormal uniform B cell population was observed however antigen expression patterns varied greatly between patients. BM from acute myeloblastic leukemia (AML) patients showed an abnormal distribution of CD34+ cells. In addition, a correlation was observed between pre-BMT cytogenetics and MPFC. Only 2 out of 8 (25%) patients with normal MPFC pre-autologous bone marrow transplantation (ABMT) relapsed (AML), while 6 out of 13 (46%) patients with abnormal pre-BMT MPFC relapsed including 2 out of 3 patients who were transplanted in clinical CR. Pre-BMT MPFC may thus be an effective tool for detection of MRD by detection of a pre-transplant MPFC abnormality.     

Bone marrow purging with mafosfamide--a critical survey

Autologous bone marrow transplantation (ABMT) is increasingly used to consolidate remissions, primarily in hematological disease. Various purging strategies have been developed to minimize the risk of reimplantation of tumor cells with the bone marrow autotransplant. Pharmacological purging with the oxazaphosphorine derivative mafosfamide has been studied extensively, and recent clinical data suggest that purging with mafosfamide may translate into superior remission duration if compared to nonpurged ABMT in acute leukemia. Chemical and experimental data relevant to mafosfamide-purging and clinical results are reviewed, with special emphasis on safety aspects.     

Autologous transplantation of a bone marrow graft manipulated by chemoseparation to eliminate residual tumor cells

An autologous bone marrow transplantation (ABMT) was performed in a 45-year-old male patient with AML in therapy-resistant first relapse including CNS-disease. The marrow graft was harvested 18 months before, at the beginning of first remission, and was subjected to an in-vitro incubation with 4-hydroperoxycyclophosphamide (4-HC) prior to cryopreservation to eliminate residual clonogenic tumor cells. After myeloablative therapy with high-dose Cyclo-phosphamide (CY), total body irradiation (TBI) and intrathecal application of Methotrexate (MTX), the manipulated marrow graft was reinfused. The myelopoietic reconstitution started already 6 days after ABMT and was completed 40 days thereafter. There is evidence for a complete remission in marrow and spinal fluid observed over a period of 4 months.     

Autologous cryopreserved platelets and prophylaxis of bleeding in autologous bone marrow transplantation

Autologous platelets were harvested and cryopreserved in eight consecutive patients elected for ablative chemotherapy and autologous bone marrow transplantation (ABMT) for solid malignancy. There was a 19% loss in platelet count after the freeze thaw and wash procedure; with an in vitro functional loss of 40-60%. No correlation could be found for individual platelet transfusions between in vitro functional tests and in vivo recovery. Six consecutive patients received a total of 16 autologous platelet transfusions in the aplastic phase of ABMT. No bleeding was observed during the study period and there was no CMV infection in the recipients. While improvement in freezing and subsequent handling is desirable, autologous cryopreserved platelets can safely be used for the prophylaxis of bleeding during aplasia in patients treated with ABMT.     

Cryopreservation of marrow, purging and autologous bone marrow transplantation in childhood

Since 1984 bone marrow from 42 children with acute lymphoblastic leukaemia, non Hodgkin's lymphoma and neuroblastoma was cryopreserved. In 5 cases (c-ALL, NHL and B type) the marrow was purged by using a cocktail of three monoclonal antibodies (VIL A1, VIB C5, VIB-E3). Up to now 13 children (ALL/10, neuroblastoma/3) were autografted (one of them after purging) after supralethal chemoradiotherapy. Except one child with early death all patients had engraftment: a level of 1.0.10(9)/l leukocytes was reached at days 10-33 (median, 19); platelet level over 60.10(9)/l at days 32-60 (median, 41). 2 children died on treatment related complications, one on infection after full haematological restitution, 2 patients alive with relapse, 8/13 alive in CCR and well.     

Early and late effects in the bone marrow of mice following 2 Gy (6 MeV) neutron irradiation

Summary Following 5 Gy gamma irradiation, residual damage in bone marrow persisted up to one year and was ascribed to genetic defects in hemopoietic stem cells (von Wangenheim et al. 1986). To see whether high LET radiation is more efficient in inducing late effects, mice were whole-body irradiated with a single dose of 2 Gy neutrons ( = 6 MeV) and femoral cellularity, CFU-S number, proliferation ability of bone marrow cells (PF) and the compartment ratio (CR), i.e. the splenic 125-iodo-deoxyuridine incorporation per transfused CFU-S were measured up to one year after the radiation insult. Within 12 weeks, femoral cellularity, PF and CR recovered to control or near-control level, whereas CFU-S numbers remained significantly below control. No further recovery was observed. On the contrary, PF and CR deteriorated again after 12 and 26 weeks, respectively. CFU-S per femur tended to decrease as well. Thus it is demonstrated that a single dose of 2 Gy 6 MeV neutrons causes significant injury in function (PF) and structure (CFU-S numbers, CR) of bone marrow which persisted up to one year. While this residual injury can be attributed to genetic defects in hemopoietic stem cells, its increasing expression is probably due to late evolving damage in microenvironmental cells. The RBE of 6 MeV neutrons for the introduction of late effects in the bone marrow is in the range of 3.     

Toxic effects of alkyl-lysophospholipids on human bone marrow and de novo leukaemias. A short overview.

Alkyl-lysophospholipids (ALPs) are reported to have an antineoplastic activity against leukaemic cells. We have tested some halogen-containing ALPs from the Central Institute of Molecular Biology (H. Brachwitz) in comparison with racemic 1-ostadecyl-2-methyl-glycero-3-phosphocholine (ET-18-OCH3) (P.G. Munder, Max-Planck-Institut für Immunobiologie, Freiburg, FRG). We found freshly dissolved ALPs to be very toxic both to human bone marrow and to leukaemic cells of patients. ALP-incubation before cryopreservation is more toxic to bone marrow (but not to AML blasts) than after cryopreservation. All experiments to test the selectivity and to establish a purging protocol should be done using 1, remission marrow including a cryopreservation step and 2, blasts of de novo leukaemias instead of cell as sensitive as HL 60 to ALP-incubation. We found direct toxicity of ALPs to be not suitable for purging lines of bone marrow from patients in remission.     

A prospective analysis of low-grade gastric malt lymphoma after Helicobacter pylori eradication

BACKGROUND: Primary gastric low-grade B-cell lymphoma of mucosa-associated lymphoid tissue (MALT lymphoma) is known to be successfully treated with anti-Helicobacter pylori (H. pylori) therapy alone. However, there are few reports on long-term results after eradication therapy. The aims of this study were to analyze the rate and the interval to reach complete remission (CR), and to assess the rate and the factors affecting recurrence of MALT lymphoma. MATERIALS AND METHODS: Between 1996 and 2003, a total of 90 H. pylori-infected patients with low-grade MALT lymphoma were included in this study. For initial staging, endoscopic ultrasonography, chest-abdomen-pelvis CT scans, and bone marrow examination were taken. All patients were made to take anti-H. pylori therapy for 14 days. Tumoral response was assessed by endoscopy every 3 months till CR and every 6 months after achieving CR. RESULTS: Among 90 treated patients, 85 (94.4%) reached CR. The median interval to CR was 3 months (range, 1-24). Seventy-nine (92.9%) patients were in CR at 12 months. Median follow-up period after CR was 45 months (range 15-109). Among 77 patients who were followed-up after CR, 8 (10.4%) patients were proved with recurrence of MALT lymphoma. Cumulative recurrence rate was 2.7, 11.5, and 12.2% at 1, 2, and 3 years. The presence of H. pylori was only a significant risk factor affecting recurrence. CONCLUSIONS: The status of H. pylori is the most important risk factor affecting recurrence. Therefore, adequate eradication regimen and accurate regular evaluation for H. pylori status are needed during follow up of primary gastric low-grade B-cell MALT lymphoma.     

Oncolytic viral purging of leukemic hematopoietic stem and progenitor cells with Myxoma virus

High-dose chemotherapy and radiation followed by autologous blood and marrow transplantation (ABMT) has been used for the treatment of certain cancers that are refractory to standard therapeutic regimes. However, a major challenge with ABMT for patients with hematologic malignancies is disease relapse, mainly due to either contamination with cancerous hematopoietic stem and progenitor cells (HSPCs) within the autograft or the persistence of residual therapy-resistant disease niches within the patient. Oncolytic viruses represent a promising therapeutic approach to prevent cancer relapse by eliminating tumor-initiating cells that contaminate the autograft. Here we summarize an ex vivo “purging” strategy with oncolytic Myxoma virus (MYXV) to remove cancer-initiating cells from patient autografts prior to transplantation. MYXV, a novel oncolytic poxvirus with potent anti-cancer properties in a variety of in vivo tumor models, can specifically eliminate cancerous stem and progenitor cells from samples obtained from acute myelogenous leukemia (AML) patients, while sparing normal CD34+ hematopoietic stem and progenitor cells capable of rescuing hematopoiesis following high dose conditioning. We propose that a broader subset of patients with intractable hematologic malignancies who have failed standard therapy could become eligible for ABMT when the treatment schema is coupled with ex vivo oncolytic therapy.     

Failure of bone marrow cryopreservation in chronic granulocytic leukemia: relation to excessive granulo-macrophagic progenitor pool

Autologous bone marrow transplantation (ABMT) in chronic granulocytic leukemia (CGL) aims at reversing the acute or acceleration phases by injection of stem cells collected during the chronic phase. This study was designed to explain an unusual rate of delayed engraftment (50%) in our experience of ABMT in CGL patients. We investigated all the factors possibly responsible for abnormal perpetuation of aplasia following infusion of cryopreserved marrow stem cells. The study of CFU-gm recovery in 41 bags of frozen marrow from 25 patients revealed an overall deficiency with a mean CFU-gm recovery of 55 +/- 38% in CGL patients versus 73 +/- 15% in the control group (p less than 0.001). Our data also showed an inverse linear relation (r = -0.40, p less than 0.05) between CFU-gm concentration and recovery after freezing. A good CFU-gm recovery (greater than or equal to = 50%) was observed in 70% of cases when the concentration was less than 3700 CFU-gm/ml as compared to 30% of cases when the concentration was over 3700 CFU-gm/ml (p less than 0.001). The lack of improvement by diluting rich CFU-gm marrows to reduce CFU-gm concentration/ml, as well as the absence of relationship between CFU-gm recovery after freezing and nucleated cells concentration, suggest a particular fragility of CGL stem cells to freezing, probably related to their excessive amplification. At the present time, we strongly recommend that the highest possible dose of progenitor cells be cryopreserved, preferably at a low concentration, in patients with CGL, and particular attention devoted to the freezing procedure in each individual patient, with numerous appropriate efficiency tests.     

Allogeneic bone marrow transplantation in children with acute lymphoblastic leukaemia in the first and second complete remission conditioned with fractionated total body irradiation and cyclophosphamide or etoposide

Patients and methodsFrom 1993 to 2001 thirty-two children underwent bone marrow transplantation (BMT) for acute lymphoblastic leukaemia (ALL) (12 in I complete remission /I CR/of high-risk/HR/ALL, and 20 in II CR after early bone marrow or combined bone marrow/organ relapse). Except for two syngeneic all others were matched sibling donor transplants. All patients (pts) were conditioned with fractionated total body irradiation (FTBI) at a total dose of 12,6 Gy, given in 8 fractions during 4 days with lung shielding (9,4 Gy) and cyclophosphamide (CY) 60 mg/kg i.v for 2 days (total dose 120 mg/kg) (n = 1 in I CR and n = 11 in II CR) or etoposide (VP) 60 mg/kg i.v (n = 11 in I CR and n = 9 in II CR). Patients in I CR were given 1,1–4,9×10⁸ nucleated cells /kg (med. 2,7×10⁸/kg), while pts in II CR 1,9–4,0×10⁸ nucleated cells/kg (med. 2,7×10⁸/kg). For graft versus host disease (GvHD) prevention cyclosporin A (CsA) 3 mg/kg/d i.v was administered alone in 22 pts (n = 9 in I CR and n = 13 in II CR) or in combination with “short” methotrexate +/− prednisone in 8 pts (n = 3 in I CR and n = 5 in II CR). Two pts transplanted with syngeneic BM received no GvHD prevention. The regimen related toxicity (RRT) was graded according to the system developed by Bearman et al. (1988).ResultsOnly mild or moderate expression of RRT was observed (GI toxicity I⁰ – 80%, II⁰ – 4%; stomatitis I⁰ – 40%, II⁰ – 20%; hepatic toxicity I⁰ – 28%; renal, bladder and cardiac toxicity I⁰ – 4%) and no transplant related deaths occurred (TRM = 0%). Among 12 pts transplanted in I CR only one child relapsed 4 months from BMT, while the remaining 11 pts are alive in continuous complete remission (CCR) with a median follow-up of 33 months (range 6 to 66 months) and 92% probability of a 5-year event free survival (pEFS). Of 20 children transplanted in II CR 6 relapsed 1–14 months from BMT (median 6,5 months). Thirteen of them remain in CCR with a median follow-up of 19.5 months (range 1 to 96 months) and with 66% probability of a 8-year EFS.Conclusions1. In children with ALL the FTBI-12,6 Gy-containing regimen is well tolerated without life-threatening toxic complications. 2. The FTBI-12,6 Gy-containing regimen demonstrates very good antileukaemic efficacy for HR-ALL in I CR, but only limited efficacy for ALL in II CR. 3. In the context of good tolerance of FTBI in a total dose of 12,6 Gy and its limited antileukaemic efficacy in children with ALL in II CR the escalation of FTBI total dose from 12,6 Gy to at least 13,2 Gy appears to be justified in those children.     

Combined ex vivo treatment with immunotoxins and mafosfamid: a novel immunochemotherapeutic approach for elimination of neoplastic T cells from autologous marrow grafts

We evaluated a novel ex vivo "purging" protocol for selective elimination of neoplastic T cells from human marrow by using a sensitive clonogenic assay. Immunotoxins (IT) were synthesized by conjugating ricin (R) to four different monoclonal antibodies (MoAb) directed against distinct markers of T cell lineage. Treatment with anti-p67-R produced effective elimination of leukemic T cells from human marrow. The cyclophosphamide congener mafosfamid (ASTA Z 7577) markedly enhanced the target cell cytotoxicity of IT and extended the final level of clonogenic kill 2 to 3 logs. Our data show that anti-p67-R in combination with mafosfamid resulted in a maximum elimination of 6.2 logs of neoplastic T cells with minimal toxicity to normal bone marrow progenitors. The efficiency of this protocol was not reduced in the presence of excess normal bone marrow cells. Similar findings were obtained by using a cocktail of four different anti-T cell IT. This approach is unique in combining both immunologic (IT) and chemical (mafosfamid) strategies for more effective ex vivo bone marrow purging in autologous bone marrow transplantation for T cell acute lymphoblastic leukemia/lymphoblastic lymphoma.     

Elimination of clonogenic tumor cells from bone marrow using methylprednisolone (MP) and etoposide VP16: an in vitro pharmacologic study

The ability to eliminate malignant cells from bone marrow (BM) while retaining sufficient numbers of normal progenitors to ensure engraftment, may well establish the future of autologous BM transplantation (ABMT) for hematologic malignancies. In this study, we describe the effects of methylprednisolone (MP) and etoposide (VP16) alone or in combination on 5 tumor cell lines (HL-60, a promyelocytic cell line; Molt-4, a T cell leukemia; Daudi, a Burkitt's lymphoma and R10/8226 and R40/8226, doxorubicin-resistant myeloma cell lines). The tumor cell kill efficiency of the drugs was assayed using the limiting dilution assay. We determined the toxic effect on progenitor cells by assaying granulocyte-macrophage colony-forming units (CFU). With a combination of MP at 10(-3) M and VP16 at 75 microM, we observed the following log reduction in tumor cell clones: HL-60, 4.695 +/- 0.001; Molt-4, 3.626 +/- 0.036; Daudi, 5.633 +/- 0.001; R10/8226, 3.052 +/- 0.544; R40/8226, 3.126 +/- 0.080. CFU recovery was 24% +/- 5%. Mixing tumor cell lines with a 20-fold excess of normal irradiated BM cells did not eliminate the inhibitory effect of the drug combination. We propose that MP and VP16 used in concert produce effective purging of malignant hematopoietic cells from BM while sparing normal progenitors needed for engraftment.     

Magnetic resonance (MR) evaluation of bone marrow in vertebral bodies.

Magnetic Resonance (MR) imaging was used to examine the hematopoietic bone marrow in the vertebral bodies of eight healthy subjects, and of 35 cancer patients who had been previously treated with radiation therapy. MR was instrumental in distinguishing viable hematopoietic tissue (red marrow) from adipose tissue (yellow marrow), whose presence reflected the extent of radiation-induced bone marrow injury. Different water content in proliferating hematopoietic tissue and adipose tissue enabled clear distinction of the two components even inside the same vertebral body. Three patterns of bone marrow viability were observed in irradiated patients: 1. Patients undergoing therapy at the time of MR study, and patients who had received low-intermediate dose several years before MR examination showed no alteration as compared with healthy controls (i.e. homogeneous presence of red marrow). 2. Patients who had received low-intermediate dose few years before MR, showed either partial re-colonization of yellow marrow or almost complete ablation of active red marrow with rare areas of re-colonization. 3. Patients who had received high dose, showed complete depletion of red marrow (fatty substitution) independently of the length of time elapsed since radiation therapy. Therefore, bone marrow recovery after radiation therapy was associate with two variables: received dose and length of time allowed for re-colonization by surviving hematopoietic tissue. In conclusion, our results provide evidence that MR can be purposively used to study composition and distribution of normal bone marrow, and to asses the extent of radiation-induced bone marrow injury; to monitor bone marrow recovery (or the lack of it); and in the general follow-up of treated cancer patients.     

Bone marrow transplantation in children with neuroblastoma

Neuroblastoma is the third most frequent malignant tumor in childhood. One-third of the patients over one year of age at the time of diagnosis suffer from the disseminated form (stage IV). Despite highly aggressive chemotherapy survival rates are poor. One hundred and eighty-seven patients with neuroblastoma stage IV have been treated according to the German protocol NB 85. The probability of disease free survival is only 15% after 70 months. Treatment strategy in our protocol includes autologous and allogeneic bone marrow transplantation (BMT) for patients with stage IV (and greater than 1 year old). Twenty-two patients were grafted (7 allogeneic and 15 autologous). The conditioning regimen consisted mainly of high-dose melphalan (180 mg/m2) and total body irradiation (TBI) (3.4 Gy). Survival rates are discussed in the context of the chemotherapy protocol. Our own experience with autologous BMT is poor, despite of different purging methods. For this reason we decided to focus on allogeneic BMT. We have grafted five patients within the last 3 years. Three of them are alive and well, on died from veno-occlusive disease 70 days after BMT, and the remaining patient, grafted from a syngeneic donor, died from relapsing tumor. The main problem in neuroblastoma stage IV is resistance to chemotherapy. Intensification of the conditioning regimen or double autografting leads to a rate of toxic deaths close to 20% (Zucker, EBMT 1987) which is not tolerable. New improvements in the conditioning regimen have to be found to increase the effect of BMT.     

Intravenous and oral demethoxydaunorubicin (NSC 256-439) in the treatment of acute leukemia and lymphoma: a pilot study

Demethoxydaunorubicin (DMDR), a new anthracycline available both for intravenous and oral administration, was given in 14 cases of leukaemia, non-Hodgkin's lymphoma (NHL) and multiple myeloma (MM) replacing either daunorubicin (DNR) or doxorubicin (DOX) in conventional chemotherapy regimes. In acute leukaemia (6 myeloblastic and 1 common lymphoblastic) there were 5 complete (CR) and 2 partial (PR) remissions; one patient, previously brought into remission with a regime including i.v. DMDR was thereafter maintained in CR with oral DMDR. Among the patients treated with the oral DMDR, 2 NHL cases were treated; 1 patient had a sustained remission of 12 months so far, with DMDR alone; another patient had a CR with a combined regime. In MM, one patient with very advanced disease treated with i.v. DMDR/CHOP did not respond, but three cases treated with oral DMDR plus other drugs showed a partial remission. Toxic effects were limited to brief episodes of nausea and vomiting in a few i.v. treated patients; a prolonged bone marrow depression was observed in one case only. No cardiotoxic effect was recorded.     

Immunological evaluation of patients with hematological malignancies receiving ambulatory cytokine-mediated immunotherapy with recombinant human interferon-α2a and interleukin-2

Summary Immunological parameters were evaluated in patients treated with cytokine-mediated immunotherapy (CMI) consisting of low doses of recombinant human interferon 2a (rIFN) and recombinant human interleukin-2 (rIL-2) administered either concomitantly or sequentially by subcutaneous self-injections in an outpatient setting. Twenty-six patients with hematological malignancies and 2 metastatic melanoma patients in a progressive stage were enrolled in this clinical trial. Of the 26 patients, 24 were at a stage of minimal residual disease, including 14 patients who had received autologous bone marrow transplantation (ABMT) 2–5 months previously, 7 chronic myelogenous leukemia (CML) and 3 acute myeloid leukemia (AML) patients. Two patients (1 CML and 1 mult. myeloma) were treated at a stage of progressive disease. Non-MHC-restricted cytotoxicity directed against natural-killer(NK)-resistant (Daudi) and NK-sensitive (K562) target cells was assessed before, during and after CMI, either in fresh peripheral blood samples (spontaneous activity) or after in vitro rIL-2 activation (induced activity). Spontaneous killing activity was low prior to treatment, but increased upon termination of treatment in 10/15 evaluated cycels. rIL-2-activated cytotoxicity in vitro was markedly elevated in 8/12 and 6/8 patients after one and two cycles, respectively, of sequential treatment, as well as in 3/8 CML and 5/6 patients after one and two cycles, respectively, of concomitant treatment Activation of the T cell mitogenic response was demonstrated in 6/9 patients after concomitant CMI, while no such effect was observed throughout a sequential treatment in lymphoma and leukemia patients after ABMT. Although a direct correlation between immune stimulation and the in vivo antitumor response cannot yet be determined, our clinical observations support a beneficial therapeutic effect in a substantial number of patients. These results indicated that the ambulatory CMI protocol of rIL-2 and rIFN could stimulate the host defense immune system and may be helpful in mediating the in vivo antitumor response in patients with minimal residual disease.     

Photosensitization of leukemic cells and normal bone marrow cells by 514 nm laser light and effects of laser light on migration inhibition and lymphokine response

In a model for ex-vivo purging of bone marrow grafts, leukemic cells and normal bone marrow cells were treated with merocyanine 540 and exposed to 514 nm laser light. With this treatment, 99.9999% of leukemic cells were killed while 55% of the normal bone marrow cells survived. The deleterious effects of laser light alone in the absence of photosensitizer were not observed as determined by cell viability, cell migration, and response of target cells to human migration inhibition factor. These results indicate that laser light induced photodynamic therapy can be useful for ex-vivo autologous bone marrow purging without regard to the deleterious effects of laser light alone.     

Late marrow recurrences in childhood acute lymphoblastic leukaemia.

Thirty children with acute lymphoblastic leukemia had a recurrence in the bone marrow after treatment was stopped electively. A second haematological remission was achieved in 27 (90%), and the median duration of remission was shortest (six months) in those relapsing within six months of stopping treatment. Four of six children relapsing over one year after stopping treatment remained in second haematological remission. Leukaemic infiltration of the central nervous system developed in four patients remaining in marrow remission. It is concluded that conventional chemotherapy is unlikely to be effective in children with acute lymphoblastic leukaemia who relapse soon after stopping treatment, that "reprophylaxis" of the central nervous system probably with long-term intrathecal chemotherapy is essential, and that some patients relapsing after prolonged unmaintained remission may achieve long-term leukaemia-free survival.     

Simultaneous development of antibody-dependent cellular cytotoxicity (ADCC) and natural killer (NK) activity in irradiated mice reconstituted with bone marrow cells

Spleen cells from irradiated, bone marrow-reconstituted mice were tested for their ability to mediate antibody-dependent cellular cytotoxicity against P815 target (ADCC-P815), ADCC against sheep red blood cells (ADCC-SRBC), and natural killer (NK) activity judged as YAC-1 lysis at different times after bone marrow reconstitution. Donor-derived ADCC-P815 effectors were found to appear in the spleens 10-12 days after bone marrow reconstitution simultaneously with the appearance of donor-derived NK cells. NK cells recently derived from bone marrow are known to express the Thy-1 antigen; the phenotype of the "early" ADCC-P815 effectors was found to be the same as that of NK cells, i.e., Thy-1+, asialo-GM1+. These data suggest that ADCC-P815 effector cells belong to the NK cell population. ADCC-SRBC, in contrast to ADCC-P815 and NK activity, was already high on Day 7 after bone marrow reconstitution. However, it was mediated partly by recipient-derived effectors. ADCC-SRBC effectors were characterized to be different from ADCC-P815 effectors.