Ex vivo bone marrow purging with oligonucleotides.
Explore the source record for details and available documents.
Animals↗
SEARCH · Search PubMed
Results for “Bone Marrow Purging”
Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.
Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.
Increased survival of normal cells during laser photodynamic therapy: implications for ex vivo autologous bone marrow purging.
Laser light-induced, dye-mediated photolysis of leukemic cells was tested in an in vitro model for its efficacy in eliminating occult tumor cells for ex vivo autologous bone marrow purging. Merocyanine 540 (MC540) was mixed with acute promyelocytic leukemia (HL-60) cells in the presence of human albumin. This cell-dye mixture was irradiated with 514 nm argon laser light. Results show that in the presence of 0.1%, 0.25% and 0.5% albumin, laser light doses of 62.4 J/cm2, 93.6 J/cm2 and 109.2 J/cm2, respectively, were required for a 5 log reduction in the survival of leukemic cells. Under identical conditions, 80% to 84% of the normal bone marrow cells and 41% of the granulocyte-macrophage colony forming cells survived. The number of surviving stromal cells was reduced (1+) compared to the untreated control (4+). Mixing of irradiated bone marrow cells with equal number of HL-60 cells did not interfere with the killing of HL-60 cells treated with MC540 and laser light. The non-specific cytotoxicity of laser light alone was less than 6% for normal bone marrow cells. These results suggest that the concentration of human albumin plays an important role in laser light-induced phototoxicity. This laser light-induced selective photolysis of leukemic cells can be used in ex vivo purging of tumor cell-contaminated bone marrow grafts to achieve very high survival rates of normal bone marrow cells and granulocyte-macrophage colony forming cells.
Albumins↗
Allogeneic versus autologous purged bone marrow transplantation for neuroblastoma: a report from the Childrens Cancer Group.
PURPOSE: We have compared the toxicity, relapse rate, and progression-free survival (PFS) of high-risk neuroblastoma patients receiving identical induction therapy and myeloablative chemotherapy plus total-body irradiation (TBI) followed by allogeneic or autologous purged bone marrow transplantation (BMT). PATIENTS AND METHODS: Fifty-six patients with high-risk neuroblastoma underwent BMT at investigator and parent option if they did not have progressive disease after induction chemotherapy with cisplatin, cyclophosphamide, doxorubicin, and etoposide. After surgery and local radiation to residual tumor, myeloablative therapy consisting of etoposide, melphalan, cisplatin, and TBI was given followed by BMT. Patients with human leukocyte antigen (HLA)-compatible siblings received allogeneic bone marrow (n = 20). The remaining patients (n = 36) received autologous bone marrow that had undergone multimodality purging and had no remaining detectable tumor cells by immunocytology. RESULTS: Four of 20 allogeneic patients had a treatment-related death, compared with three of 36 autologous patients (P = .21). The relapse rate among allogeneic BMT patients was 69%, compared with 46% for autologous BMT patients (P = .14). The estimated PFS rates 4 years after BMT were 25% for allogeneic BMT patients and 49% for autologous BMT patients (P = .051). CONCLUSION: Overall outcome for patients with neuroblastoma given this same induction therapy followed by autologous purged marrow was similar to that with allogeneic marrow, although bias in patients selection cannot be excluded in a nonrandomized comparison.
Adolescent↗
Normal and leukemic hematopoietic cells manifest differential sensitivity to inhibitory effects of c-myb antisense oligodeoxynucleotides: an in vitro study relevant to bone marrow purging.
The c-myb protooncogene is preferentially expressed in hematopoietic cells, and its encoded protein, Myb, is required for hematopoietic cell proliferation. To analyze the relative Myb dependence of normal and leukemic human hematopoietic progenitor cells, normal bone marrow cells, several types of leukemic blast cells, and 1:1 mixtures of normal and leukemic cells were cultured in the presence of c-myb sense or antisense oligodeoxynucleotides; cell viability and cloning efficiency were then assessed. c-myb sense oligomers had negligible effects on normal and leukemic cells. In contrast, c-myb antisense oligomers strongly inhibited or completely abolished clonogenic growth of a T-cell leukemia line, 78% (18 of 23) of primary acute myelogenous leukemia cases examined, and 4 of 5 primary chronic myelogenous leukemia (CML) cases in blast crisis. In three of the latter patients, polymerase chain reaction analysis of a 1:1 mixture of c-myb antisense-treated normal and CML cells revealed a complete absence of bcr-abl expression, suggesting that the CML clonogenic units had been completely eliminated from the cultures. At antisense doses that inhibited leukemic cell growth, normal hematopoietic progenitor cells survived. Thus, normal and leukemic hematopoietic cells show differential sensitivity to the toxic effects of c-myb antisense DNA. Perturbation of c-myb function with antisense oligodeoxynucleotides might eventually form the basis for a molecular approach to leukemia therapy, perhaps most immediately as ex vivo bone marrow purging agents.
Base Sequence↗
Autologous transplantation of bone marrow purged in vitro with anti-CD7-(WT1-) ricin A immunotoxin in T-cell lymphoblastic leukemia and lymphoma.
Seven patients with high-risk acute T-cell lymphoblastic leukemia (T-ALL) and six with T cell lymphoma (T-LL) were treated with autologous bone marrow transplantation (ABMT) after in vitro purging of their bone marrow with WT1 (CD7)-ricin A-chain immunotoxin. CD7 expression on the tumor cells showed large variations between the individual patients and was highly related to the specific cytotoxicity of WT1-ricin A. Incubation of bone marrow with up to 10(-8)mol/L WT1-ricin A in the presence of 6 mmol/L NH4Cl did not compromise the growth potential of the hematopoietic progenitors CFU-GM, CFU-GEMM, and BFU-E. Hematologic engraftment (greater than 10(9) leukocytes/L) occurred within a normal time period (median, 17 days). Seven patients are alive and in complete remission (CR) at 48+, 44+, 40+, 26+, 11+, 7+, and 6+ months after ABMT. Four patients relapsed within 6 months after ABMT. Two of them had the lowest CD7 expression on their tumor cells, the other two were transplanted in CR2 and CR3. Two patients died from transplantation related infections. The immunologic reconstitution was delayed, although the numbers of T cells reached normal levels within 1 month. The number of CD7+ cells remained low up to 1 year after transplantation. The T4/T8-ratio was decreased for at least 6 months. The T-cell response to mitogens recovered to normal levels after 1 year. This study shows that ABMT with WT1-ricin A purged bone marrow in high-risk T-cell malignancies results in a complete hematopoietic and a delayed immunologic reconstitution. The actuarial relapse free survival is 61% at 3 years.
Adolescent↗
Value of autologous stem cell transplantation with purged bone marrow as first-line therapy for follicular lymphoma with high tumor burden: a GOELAMS phase II study.
This prospective phase II study was undertaken to evaluate the efficacy and toxicity of early intensive therapy followed by purged autologous bone marrow transplantation (ABMT) in patients with follicular lymphoma with high tumor burden. All patients received the VCAP regimen (vindesine, cyclophosphamide, doxorubicin and prednisone) as conventional chemotherapy and DHAP as second-line therapy. Twenty-nine consecutive patients were included in the study. Twenty-seven patients were grafted, seven in first complete remission (CR) and 20 in first partial remission (PR). Preparative therapy consisted of cyclophosphamide and total body irradiation (TBI) in all the patients. With a median follow-up of 6 years, the actuarial overall survival is 64% and the actuarial event-free survival is 55%. Two treatment-related early deaths were observed. Eleven patients were informative for serial PCR analysis of minimal residual disease after ABMT: two relapsed, four remained disease-free with PCR positivity and five were disease-free with PCR negativity. These encouraging results lay the basis of future prospective randomized trials comparing autologous stem cell transplantation as front-line treatment with conventional chemotherapy for patients with bad prognostic factors.
Adult↗
Monoclonal antibody-purged bone marrow transplantation therapy for multiple myeloma.
This report describes the clinical characteristics, treatment associated toxicity, and follow-up of fifty-eight patients with plasma cell--dyscrasias treated with high dose chemotherapy and total body irradiation (TBI) at a single institution. Following TBI, 36 patients received anti-B cell monoclonal antibody (MoAb)-treated autologous bone marrow, 21 patients received anti-CD6 cell MoAb-treated allogeneic bone marrow to deplete T cells, and one patient received unpurged bone marrow from a syngeneic donor. Evaluation after high dose chemotherapy and bone marrow transplantation (BMT) demonstrated 26 complete responses (CR), 26 partial responses (PR), 2 non-responders, 1 not yet evaluated, and three toxic deaths. Fourteen of 36 patients who underwent autologous BMT are alive free from progression at 18 (range 5 to 68) months post transplant (post-BMT); of these, 11 remain in continuous complete response at 16 (range 5 to 68) months post-BMT. Seven of 21 patients who underwent allogeneic BMT are alive free from progression at 30 (range 4 to 44) months post-BMT; of these, three patients remain in continuous complete response at 43 (range 33 to 45) months post-BMT. These data suggest that high dose chemotherapy with TBI followed by MoAb purged BM can be performed with acceptable toxicity and high tumor response rates.
Adult↗
Potential of phenylalanine methylester as a bone marrow purging agent.
Phenylalanine methylester (PME), a lysosomotropic compound can be used to deplete monocytes and myeloid cells from peripheral blood and bone marrow (BM). The potential of PME for purging leukemic cells from BM was investigated using U937 and HL-60 cell lines as models. Optimal purging conditions for U937 cells were determined using an MTT assay (3-4, 5-dimethylthiazol-2, 5-diphenyl tetrazolium biomide; Sigma). Elimination of U937 cells was time-, temperature-, and dose-dependent. PME activity was optimal at 37 degrees C for 45 minutes. Depletion of U937 was > 2.8 logs for 50 mmol/L PME. Compared with another purging agent, 100 micrograms/mL 4-hydroperoxycyclophosphamide had activity comparable to 40 mmol/L PME. HL-60 cells were even more sensitive to PME than U937 cells. To support observations made with the MTT assay, clonogenic assays were performed. PME, 50 mmol/L at 37 degrees C resulted in total depletion (> 5 logs) of U937 colonies. Progressive depletion of normal progenitor cells occurred when BM was incubated with PME at concentrations from 5 to 100 mmol/L. At 37 degrees C, 50 mmol/L PME reduced colony-forming units-granulocyte-macrophage and burst-forming units-erythroid (BFU-E) recovery by 98%. Recombinant human mast cell factor augmented BFU-E after PME treatment but had no effect on HL-60 or U937. These studies suggest that PME deserves further study as an agent for ex vivo marrow purging.
Bone Marrow↗
Combinations of 4-hydroperoxycyclophosphamide (4-HC) and cisplatin for bone marrow purging in autologous marrow transplantation: an update.
We are studying the usefulness of combinations of 4-HC and cisplatin as a potential purging regimen for autologous bone marrow transplantation. In all of our studies, in vitro cytotoxicity was determined by clonogenic assay, and drug interaction was quantitated using the multiple drug-effect analysis method. The cells were incubated for one hour (4-HC) and/or 4 hours (cisplatin). We found that the drugs in combination had cytotoxic synergism against human leukemia cell lines (K-562 and Raji). The synergism was sequence-dependent (cells must be exposed to 4-HC first), was present at various molar ratios of the drugs, and most pronounced at high levels of cell kill. We also found that the drugs had cytotoxic synergism against normal human marrow progenitors (CFU-GM). However, the leukemic cells were approximately 55 times more sensitive to the combination than CFU-GM. In a murine system, the drugs were synergistic against L1210 leukemia cells and normal murine CFU-GM, but L1210 cells were at least 130 times more sensitive to the combination than CFU-GM. To determine the ability of L1210 cells to grow in vivo after exposure to the drugs, BDF1 mice were injected with 2 x 10(4) cells which had been incubated with 4-HC and/or cisplatin. The survival time of untreated controls was 13 +/- 2.8 days. For treated groups, the cure rates after 50 days of observation were 33% (4-HC, 40 uM), 0% (cisplatin, 8 uM), and 100% (4-HC + cisplatin). Finally, at concentrations resulting in equivalent toxicity to marrow CFU-GM, cisplatin seemed to be more toxic to murine spleen blast colony forming cells (CFC-BC) than 4-HC. The drugs in combination appeared to have at least additive toxicities against CFC-BC.
Animals↗
[Massive chemotherapy and purged bone marrow autograft in severe neuroblastoma. Preliminary results apropos of 5 cases].
Massive chemotherapy with in vitro purged autologous bone marrow transplantation has benefited from better understanding of massive chemotherapy for solid tumors, and better search for bone marrow involvement; it may now be used for treatment of poor prognosis neuroblastomas. Authors report preliminary results of 5 cases (4 IV and 1 p III b stage of TNM classification) completely treated in one institution (Centre Léon-Bérard, Lyon, France). Three patients are in complete remission, and 2 have relapsed (median follow up: 18 months). Authors comment on these results.
Adrenal Gland Neoplasms↗
The effects of acetaldophosphamide, a novel stable aldophosphamide analogue, on normal human and leukemic progenitor cells in vitro: implications for use in bone marrow purging.
Acetaldophosphamide (A-ALD), a novel in vitro active and stable derivative of aldophosphamide, kills human bone marrow-derived granulocyte-macrophage colony-forming cells (GM-CFC) independent of the cell cycle. The surviving fraction of GM-CFC is an exponential function of the drug concentration and time of exposure. Variation of marrow light-density cell concentration between 2 x 10(6) and 10 x 10(6)/ml does not significantly influence its GM-CFC toxicity. Marrow depleted of GM-CFC by A-ALD subsequently generates GM-CFC when grown in suspension cultures. During the early period after treatment with A-ALD the number of surviving GM-CFC (size of surviving GM-CFC compartment) does influence the speed of the GM-CFC repopulation in suspension cultures. The importance of the number of surviving GM-CFCs for the growth and maintenance of GM-CFC population in such suspension cultures diminishes with time. No significant differences are observed after 2 wk, indicating that the ancestor stem cell population and its regenerative potential responsible for in vitro hematopoiesis have not been significantly affected by the drug treatment. A-ALD-treated progenitor cells retain their ability to integrate with the previously established marrow stromal cell layer and generate GM-CFC within this layer to an extent comparable to that of untreated marrow cells. The effect of A-ALD on human hematopoiesis is comparable to that of 4-hydroperoxycyclophosphamide. Its advantage over 4-hydroperoxycyclophosphamide is a greater stability in vitro. It has sparing effect on GM-CFC ancestor cells. Its toxicity to myeloid leukemia cell line (KBM-3)-derived clonogeneic cells is higher than to the GM-CFC. It is similar in doxorubicin-sensitive (KBM-3) and -resistant (KBM-3/DOX) leukemic cells. Thus, A-ALD appears to be a promising drug for in vitro purging of bone marrow cells.
Bone Marrow↗
Bone marrow purging, 4-hydroperoxycyclophosphamide, and the FDA.
Explore the source record for details and available documents.
Acute Disease↗
Bone marrow purging with oligodeoxynucleotides.
Explore the source record for details and available documents.
Bone Marrow Purging↗
Quantitation of GD2 synthase mRNA by real-time reverse transcription-polymerase chain reaction: utility in bone marrow purging of neuroblastoma by anti-GD2 antibody 3F8.
BACKGROUND: Antigen ganglioside GD2 is expressed abundantly on neuroblastoma (NB) cells. Anti-GD2 monoclonal antibody (MoAb) 3F8 kills NB cells by complement-dependent cytotoxicity and antibody-dependent cellular cytotoxicity. Its utility in bone marrow (BM) purging is evaluated by a real-time reverse transcription-polymerase chain reaction (RT-PCR) assay to quantify the mRNA of GD2 synthase, the key enzyme in GD2 synthesis. METHODS: From 1990 to 1993, 10 patients with relapsed/refractory Stage 4 NB participated in a pilot study. In these patients, MoAb 3F8 was used to purge tumor cells from harvested BM that had 5% or less tumor content by immunofluorescence (IF). Subsequently, 31 Stage 4 NB patients who underwent treatment on the N7 protocol (1994-1999) had their BM, which was in remission, purged by 3F8 before (131)I-3F8 myeloablative radioimmunotherapy. GD2-positive tumor cells before and after purging were quantified by real-time quantitative RT-PCR of GD2 synthase. RESULTS: GD2 positivity by IF was found before purging in six of eight patients in the pilot study. Five of six patients became negative postpurging. Of 31 patients on the N7 protocol, the more sensitive real-time quantitative RT-PCR detected GD2 synthase mRNA in the BM samples of 7 patients even though the prepurge BM samples were negative by histology and IF. Six of the seven BM samples became negative after 3F8 purging. Marker positivity before purging was statistically significant in predicting overall survival (P = 0.04), but not progression-free survival (P = 0.1). In vitro hematopoietic stem cell recovery and the median time to engraftment were acceptable. CONCLUSION: Tumor cell depletion quantified by real-time RT-PCR demonstrated efficacy of MoAb 3F8 in BM purging.
Antibodies, Monoclonal↗
Generation of human natural killer cells from pharmacologically purged bone marrow.
Human natural killer (NK) cells play an important role in first-line defence against primary and metastatic tumours. Their stimulation after autologous bone marrow transplantation (ABMT) may be useful to eradicate residual tumour cells. Human NK cells originate in bone marrow (BM) but little is known about their progenitors and lineage development. We studied NK cell ontogeny from BM progenitors obtained by 'purging' normal BM with 4-hydroperoxycyclophosphamide (4HC); this agent is known to destroy all but the most primitive BM progenitors. NK cells, defined by their phenotypic markers and cytolytic activity, could be generated from 4HC-treated BM cells during in vitro cultures over stromal BM feeder layers and in suspension cultures containing a mixture of soluble cytokines; Interleukine 2 appears to be essential for the full development of this population. A lag period exists before NK cells can be found in significant numbers in culture; this suggests that a delay in initiation of immunotherapy after ABMT ought to be considered, particularly when using purged marrow.
Antigens, CD↗
The effect of tumor necrosis factor on normal human hematopoietic progenitors.
Tumor necrosis factor-alpha (TNF-alpha), a product of activated macrophages that is cytotoxic to tumor cells, could be used to purge tumor cells from bone marrow before autologous bone marrow transplantation for hematologic malignancies and/or solid tumors. To determine whether exposure to TNF-alpha would have an inhibitory effect on hematopoietic progenitors, we incubated normal human bone marrow with a wide range of concentrations of recombinant human TNF. In order to mimic the conditions that would be used in bone marrow purging, bone marrow cell suspensions were incubated with TNF in doses ranging from 500 to 100,000 U/ml for 24 hours, and were assayed for colony formation in agar. We noted a dose-dependent inhibition of total colony-forming units (CFU) at days 7 and 14, with 50% inhibition occurring at 60,000 U/ml of TNF. TNF exerted a differential effect on CFU so that colony formation by erythroid (CFU-E), multipotential (CFU-GEMM), and macrophage (CFU-M) progenitors was suppressed to a greater extent than that by granulocyte progenitors (CFU-G). However, even after preincubation with TNF at high doses such as 100,000 U/ml, the inhibitory effects of TNF could be abolished by washing cells before culturing. This study demonstrates that hematopoietic precursors survive treatment with TNF at doses that have been shown to be cytotoxic to tumor cells. Although TNF has a significant inhibitory effect on the growth of erythroid, multipotential, and macrophage progenitors in vitro, this effect depends on continuous exposure to TNF for more than 24 hours. Thus, TNF may be useful as a bone marrow purging agent against tumor cells, with relative sparing of normal marrow elements.
Colony-Forming Units Assay↗
Pharmacologic bone marrow purging: is there any place for etoposide? In vitro comparison with mafosfamide.
Residual leukemic cells in a bone marrow graft may increase the risk of relapse after autotransplantation. We have compared the efficacy of etoposide with that of mafosfamide, which has been used mainly for purging in acute leukemias. First, we examined the effects of VP-16 and ASTA-Z on the normal hematopoietic progenitors and on the erythroleukemic cell K562. Subsequently, we evaluated purging activity in cocultures using two different ratios of leukemic contamination. The most effective drug concentrations in inhibiting 100% of K562 growth were 50 micrograms/ml of ASTA-Z and 70 micrograms of VP-16. Residual growth of normal colony-forming units-granulocyte-macrophage (CFU-GM) was 4.8% with VP-16 and 32.5% with ASTA-Z. In treated cocultures, ASTA-Z produced a higher inhibition of the K562 line than VP-16 at both levels of leukemic cell contamination. At 0.5% contamination, VP-16 showed higher toxicity toward normal hematopoietic progenitors than ASTA-Z. At the 5% contamination level, VP-16 completely inhibited colony formation, whereas ASTA-Z spared some normal progenitor cells (21.2%). In conclusion, in our experimental model, VP-16 did not show improved efficacy over ASTA-Z in killing leukemic cells and in sparing normal progenitors.
Antineoplastic Agents↗