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Biomedical subjects

D Hoelzer

Publications and source records attributed to D Hoelzer.

At least 181 records · Page 10Linked to original sources
PubMed1995

High-dose cytosine arabinoside and daunorubicin postremission therapy in adults with de novo acute myeloid leukemia. Long-term follow-up of a prospective multicenter trial.

A total of 149 consecutive de novo AML patients aged 50 years or less (median age = 37 years) were enrolled in this prospective multicenter trial initiated in May 1985. All patients received the same induction and early consolidation therapy with daunorubicin (DNR), cytosine arabinoside (Ara-C), and etoposide (DAV). High-dose Ara-C/DNR therapy included Ara-C at 3 g/m2, in 12 doses (HD-Ara-C/DNR I) and eight doses (HD-Ara-C/DNR II), followed by DNR 30 mg/m2 for 3 days. A complete remission (CR) was achieved in 104 (70%) patients; 61 complete responders received at least one cycle with HD-Ara-C/DNR. If those patients who were transplanted in first CR (n = 26), were not considered, the median relapse-free-survival (MRFS) of the remaining 78 patients was 15 months, with a probability of relapse-free survival (RFS) at 116 months of 30% (95% CI, 20-40%) after a median follow-up of 95 months. The MRFS of the HD-Ara-C/DNR consolidated patients was 25 months, with a probability of RFS at 116 months of 37% (95% CI, 24-50%). If all patients who were transplanted (n = 44) were not considered, the median survival time (MST) was 18 months with a probability of being alive at 118 months of 24% (95% CI, 16-33%). MST of the HD-Ara-C/DNR consolidated patients was 58 months with a survival probability of 46% (95% CI, 31-60%) at 118 months. Prognostic factor analysis did not reveal any significant influence of age, sex, FAB subtype, white blood cell count, hemoglobin level, thrombocyte count, LDH, or response to the first induction course on RFS of the HD-Ara-C/DNR consolidated patients. In summary, HD-Ara-C/DNR consolidation can improve the long-term outcome of a subgroup of de novo AML patients. Further improvement of the outcome seems to depend on the identification of patients with an inferior outcome under that strategy who might benefit from alternative treatment strategies.

Adolescent↗
PubMed1995

Bone marrow-derived T-cell clones obtained from untreated acute myelocytic leukemia exhibit blast directed autologous cytotoxicity.

The eradication of minimal residual blast populations by activation of autologous cytotoxic cells with interleukin 2 (IL-2) is a new promising tool in the treatment of acute myelocytic leukemia (AML). However, the immunological effector cells are not yet clearly defined. The present study was designed to investigate the presence of cytotoxic precursor cells in active AML and to identify phenotypical and functional characteristics of autologous anti-leukemic cytotoxic effector cells. For this purpose, mononuclear cells (MNC) containing at least 70% leukemic blasts were isolated from bone marrow of untreated AML and cultured in the presence of 3000 IU/ml recombinant IL-2 (rIL-2) for 6-8 weeks. Under these conditions, T-cells were selected in the bone marrow cultures and overgrew the leukemic blasts. The resulting T-cell populations were cloned by limiting dilution and the clones obtained were characterized for their phenotypical and functional patterns. Totally, cloning resulted in 68 clones and a few cell lines. The clonality was verified by RT PCR analysis of TCR V beta gene expression. All clones obtained stained positive for CD2, CD3, DR and CD56. The vast majority (68%) of T-cell clones/lines was CD4+, a few clones expressed CD8 (19%) or CD4 and CD8, and four clones were of TCR gamma delta origin. Seven of 15 clones tested, including three CD4+, two CD8+ and two TCR gamma delta(+)-clones were found to be cytotoxic against autologous leukemic blast cells. All except one clone expressed oncolytic activities against allogeneic blasts too. One of the TCR gamma delta(+)-clones demonstrated NK activity by lysis of K562 targets. The majority of the T-cell-clones released IL-2, IL-8, TNF-alpha, GM-CSF but only a few IFN gamma and expressed high levels of mRNA for IL-2, TGF-beta and IL-10. None of the clones was found to produce IL-3, IL-4, IL-7 and TNF-beta. The data provide evidence of the existence of T-cell precursors in untreated AML bone marrow differentiating to cytotoxic cells with activity against autologous and allogeneic AML blast cells.

Bone Marrow Cells↗
PubMed1995

AML blasts variably express interleukin 2 receptor alpha, beta or gamma chains without measurable effects on proliferation, cytokine message expression or surface expression of adhesion molecules upon stimulation with interleukin 2.

Preliminary clinical studies including interleukin 2 (IL-2) in chemotherapy strategies for treatment of acute myeloblastic leukemia (AML) suggest that IL-2 may improve the disease-free survival of the patients. Because of reports showing interleukin 2 receptor expression on AML blasts, it is important to know whether IL-2 may directly influence these leukemic cells. In initial studies using flow cytometry to analyze surface expression of interleukin 2 receptors (IL-2R), we found expression of the IL-2R alpha chain on blast cells in 26% and of the IL-2R beta-chain in 81% of the patients. To confirm these results at the transcriptional level, we studied the expression of RNA of the IL-2R alpha, beta, and gamma chains by RT-PCR in the bone marrow of 38 newly diagnosed patients with AML and in three AML-derived cell lines. RNA of the alpha chain was detectable in 11/38 patients, the beta chain in 10/38 patients and the gamma chain in 30/38 patients with AML. Blast cells obtained from colonies growing in semisolid media expressed mRNA for IL-2R. RNA for all three IL-2R chains was expressed in lines KG1, HEL 92.1.7 and K562. In comparison with unstimulated cell lines, incubation of the three lines with various amounts of IL-2 over 3 and 14 days did not increase their growth or change message expression of IL-2R, IL-10, and TGF-beta and surface expression of the adhesion molecules CD 11, CD 18, CD 29 and CD 54. In conclusion, despite expression of IL-2 receptors, AML blasts do not respond to IL-2 by proliferation, message expression for various cytokine genes and surface expression of cellular adhesion molecules.

Base Sequence↗
PubMed1995

Recombinant human granulocyte-macrophage colony-stimulating factor after combined chemotherapy in high-grade non-Hodgkin's lymphoma--a randomised pilot study.

High-grade non-Hodgkin's lymphomas (NHL) can potentially be cured with combination chemotherapy, although the optimum schedules still have to be defined. Clinical trials with intensive chemotherapy are predominantly limited by myelosuppression. Here, haematopoetic growth factors open up the possibility of reducing chemotherapy-associated toxicities. In this randomised pilot study, we investigated the effects of a recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) following combined chemotherapy with vincristine, doxorubicin, cyclophosphamide, prednisone and etoposide (VACPE). A total of 35 patients with high-grade NHLs were randomised to receive either rhGM-CSF or placebo during the first two chemotherapy cycles and rhGM-CSF for all following cycles. rhGM-CSF was administered at a dosage of 5 micrograms/kg for 10 days or until neutrophils were > 1/nl following chemotherapy. The analyses revealed a significant reduction of neutropenia and duration of neutropenia in the rhGM-CSF group. Adverse events were rare and generally mild apart from one anaphylactoid reaction. No effects of rhGM-CSF were observed concerning the platelet nadir or duration of thrombocytopenia. The benefit of rhGM-CSF for response induction and survival via rhGM-CSF-supported dose intensification remains to be determined.

Adolescent↗
PubMed1995

Vincristine, doxorubicin, cyclophosphamide, prednisone and etoposide (VACPE) in high-grade non-Hodgkin's lymphoma--a multicenter phase II study.

BACKGROUND: Patients with high-grade non-Hodgkin's lymphoma (NHL) can potentially be cured by intensive chemotherapy. However, many patients still die of their disease, which underscores the need to define patient groups with different long-term prognoses and for more effective and possibly risk-adapted treatment approaches. PATIENTS AND METHODS: In this phase II study we investigated the feasibility and efficacy of a polychemotherapy consisting of 2 mg vincristine (V) on day 1, 25 mg/m2 doxorubicin (A) days 1-3, 800 mg cyclophosphamide (C) day 1, 60 mg/m2 prednisone (P) days 1-7 and 120 mg/m2 etoposide (E) days 1-3. This cycle (VACPE) was repeated on day 22 for up to 5 cycles in stages I-III and 6 cycles in stage IV, respectively, followed by consolidating radiotherapy in 38/73 patients. A total of 75 patients with high-grade NHLs according to the Kiel classification were eligible, and 73 patients are evaluable for response. The predominant histological subtypes were centroblastic, pleomorphic T-cell and large-cell anaplastic lymphomas, 60% of the patients presented with stage III/IV, 55% with a poor performance status (ECOG > or = 2), 53% with B symptoms and 60% with a LDH level >200 U/l. RESULTS: 57/73 patients achieved CR (78%), and the overall response rate (CR-PR) was 95%. The median observation time is 40 months (10+-74+). The 1-, 3- and 5-year overall survivals for the entire VACPE group were 79%, 64% and 61%, respectively. Forty-one patients are in ongoing CR with a continuous complete remission rate (CCR) of 67%. Fourteen of the 16 patients who relapsed (88%) did so within the first 24 months. The predicted 1-, 3- and 5-year DFS for those patients who achieved CR is 83%, 67% and 67%, respectively. The early mortality was 3/73 (4.1%). In patients with reduced performance status the overall survival (OS) (ECOG > or = 2) was significantly reduced, with a predicted 1-, 3- and 5-year survival of 62%, 49% and 49% versus 100%, 84% and 77% in patients with favorable performance status, respectively (p = 0.001). The predicted overall survival in stages III/IV is worse than in early stages with a 1-, 3- and 5-year probability of 73%, 52% and 52% versus 90%, 86% and 78%, respectively (p = 0.02). Comparison of patient groups with cumulative risk factors shows a significant decrease in overall survival. Especially in patients with 0-2 risk factors versus those presenting with >2 risk factors, there is a significantly better 3- and 5-year survival (p = 0.002). In contrast to overall survival, there were no differences between the listed risk groups concerning the disease-free survival of complete responders. CONCLUSION: In conclusion, the VACPE regime is feasible and effective in high-grade NHLs and may also be administered on an outpatient basis. Despite encouraging data, however, a prospective randomized trial is warranted to define a possible superiority to standard CHOP. However, this regimen may be the basis for further randomized and risk adapted innovative approaches for high-grade NHLs.

Adolescent↗
PubMed1995

The Wilms' tumor gene is frequently expressed in acute myeloblastic leukemias and may provide a marker for residual blast cells detectable by PCR.

BACKGROUND: The tumor suppressor gene wt-1 was isolated by cytogenetic deletion analysis of patients with Wilms' tumor (wt-1). This gene encodes for a zinc finger DNA-binding protein with transcription-repressing properties. During normal ontogenesis it is expressed in a time- and tissue-dependent manner mainly in the kidneys and gonads. Recently, the expression of wt-1 in acute leukemias (AL) was reported. Here we investigated the prognostic potential of wt-1 mRNA expression during the course of the disease using the PCR technique. PATIENTS AND METHODS: Blast cells from 83 patients with newly diagnosed AML and 20 AML patients during follow-up in complete remission were analyzed for wt-1 mRNA expression. Peripheral blood mononuclear cells (PBMNC) and bone marrow (BM) from healthy persons (n = 13) and sorted CD34-positive cells from normal donors (n = 4) were used as negative controls. RESULTS: Wt-1-specific m-RNA was detectable in 67/83 (81%) patients with AML. Normal donors did not express wt-1 m-RNA but in 1/4 sorted CD34+ cell samples a weak amplified product was observed. After achieving cytological CR 14/20 studied patients lost wt-1 expression. In 7/8 patients in morphological CR the reappearance of wt-1 expression preceded relapse of the disease, in 1/8 patients wt-1 remained positive in CR. Response to therapy, disease-free survival, overall survival and FAB-subtype did not correlate with wt-1 m-RNA expression in newly diagnosed AML before therapy. CONCLUSIONS: In the majority of acute leukemias wt-1 is expressed and probably blast cell-associated, at least in levels detectable by PCR. Wt-1 mRNA was detectable in bone marrow cells of AML patients in clinical CR. The results strongly suggest that the persistence or reappearance of wt-1 predicts relapse of the disease prior to morphological relapse.

Adult↗
PubMed1995

Granulocyte colony-stimulating factor and granulocyte-macrophage colony-stimulating factor in the treatment of myeloid leukemia.

Hematopoietic growth factors primarily used in patients with acute myelogenous leukemia after chemotherapy could reduce significantly the neutrophil recovery time in all patients. In high-risk acute myelogenous leukemia, trials also reported a reduction in the incidence of documented infections and early mortality rate. Thus in elderly patients with acute myelogenous leukemia and in high-risk patients with acute myelogenous leukemia the use of hematopoietic growth factors seems justified. Whether the rate of infections, particularly of life-threatening fungal infections, can be reduced by granulocyte colony-stimulating factor or granulocyte-macrophage colony-stimulating factor after chemotherapy cannot be known without larger studies. A novel strategy in the treatment of acute myelogenous leukemia is the attempt to increase the growth fraction of clonal leukemic cells prior to administration of chemotherapeutic agents by the administration of hematopoietic growth factors. Evidence shows that hematopoietic growth factors enhance anti-leukemic activity of cytosine arabinoside against leukemic cells by recruitment of leukemic cells into cell cycle, an increase of intracellular cytosine arabinoside triphosphate:deoxcytidine 5' triphosphate pool ratios, or by an enhanced cytosine arabinoside incorporation into the DNA of acute myelogenous leukemia blasts. Whether these mechanisms lead to an increase in the complete remission rate and eventually to an improvement in survival must be answered in ongoing larger acute myelogenous leukemia trials using granulocyte colony-stimulating factor or granulocyte-macrophage colony-stimulating factor in such a setting.

Acute Disease↗
PubMed1995

Changes in erythroid progenitor cell and accessory cell compartments in patients with myelodysplastic syndromes during treatment with all-trans retinoic acid and haemopoietic growth factors.

Differentiation induction therapy is used in myelodysplastic syndromes (MDS) to improve maturation defects and to restore impaired function of malignant cells. To this end, 18 patients with MDS received either a combination therapy consisting in study 1 of all-trans retinoic acid (ATRA) and granulocyte-colony stimulating factor (G-CSF), or in study 2 of a combination with ATRA, G-CSF, erythropoietin (Epo) and tocopherol. The ANC increased in 19/20 patients in both studies, whereas an increase in haemoglobin concentration, platelet counts or reduction of transfusion requirement was seen in only 8/20 patients, correlating strongly with good BFU-E growth (P < 0.001). To assess the role of accessory cells in the modulation of the haemopoietic response to treatment, we analysed the capacity of peripheral blood monocytes to secrete cytokines (IL-1 beta, IL-6, IL-8, TNF alpha). Secretion of all cytokines was significantly reduced before therapy when compared with healthy controls, but increased during therapy, reaching normal levels for IL-8. These data indicate that a combination therapy with ATRA and cytokines improves impaired cytokine secretion from monocytes and induces a multilineage clinical response in a subgroup of MDS patients characterized by an almost intact erythroid compartment. In contrast, induction of TNF alpha might be responsible for treatment failure.

Adult↗
PubMed1995

Interleukin-2 bolus infusion as late consolidation therapy in 2nd remission of acute myeloblastic leukemia.

The activation of autologous cytotoxic cells by interleukin-2 (IL-2) may be a promising tool for elimination of minimal residual blast populations in patients with acute myelocytic leukemia (AML) to prolong disease-free survival. Here, we report the results of a phase II study using IL-2 for consolidation therapy in patients with second remission of de novo AML. All patients in 1st relapse of AML received a uniform induction therapy consisting of intermediate high-dose AraC (iHDAraC) 2 x 600 mg/m2 d1-4 and VP-16 100 mg/m2 d1-7. Patients achieving 2nd remission were treated with 4 cycles recombinant IL-2 (rIL-2) 9 x 10(6) IU/m2 administered on d1-5 and 8-12/cycle as 1h infusion every six weeks. In 37/66 (56%) evaluable patients, complete remission (CR) was achieved. So far, 21/37 patients (4 after additional autologous bone marrow transplantation (ABMT) received rIL-2 consolidation. Three patients are too early for evaluation, 4 received allogeneic BMT, 6 relapsed before IL-2 was scheduled and 4 refused treatment with rIL-2. The median disease-free survival (DFS) was 11 (4-49+) months. Up to now, in 5/21 (24%) patients the duration of 2nd remission exceeded that of 1st remission 7/21 (33%) are in ongoing 2nd remission (7+ to 49+ months). The side effects of rIL-2 were generally moderate and manageable. Only in two patients, previously treated with ABMT, severe side effects occurred; septicaemia and pneumonia in one patient and desquamative erythrodermia in the second one. In accordance with other studies rebound lymphocytosis with a marked increase of CD56(+)-cells and release of secondary cytokines such as TNF-alpha, IFN-gamma and IL-6 was observed. The schedule is feasible and the data suggest a possible benefit for DFS, which, however has to be confirmed by randomized trials.

Adult↗
PubMed1995

Clinical experience with fludarabine and its immunosuppressive effects in pretreated chronic lymphocytic leukemias and low-grade lymphomas.

Fludarabine monophosphate (FAMP) is a new adenine nucleoside analogue with a promising efficacy in B-cell chronic lymphocytic leukemia (B-CLL) and low-grade non-Hodgkin lymphomas (NHLs). Here, the clinical experience and side effects with FAMP are reported in 77 patients with pretreated CLL (59 B-CLL, 2 T-CLL) and low-grade NHLs (9 immunocytic lymphomas including 5 Waldenström's macroglobulinaemia, 2 centrocytic (cc) and 5 centroblastic-centrocytic (cb-cc) NHLs). 70/77 patients are evaluable for response. All except 8 patients were pretreated with one to four different regimens and had progressive disease. FAMP was administered at a dosage of 25 mg/m2 daily for 5 days as 30 minute infusion every fifth week. Partial (PR) or complete remission (CR) was achieved in 38/56 (68%) and 3/56 (5%) of evaluable patients with CLL, respectively. In 7/8 (1 x CR, 6 x PR) evaluable patients with immunocytic lymphoma and in 3/6 (3 x PR) patients with cc or cb-cc lymphoma remissions were obtained. The probability of progression-free survival was 66% and the event-free survival was 25% and 22% at 12 and 18 months. The median progression-free survival until relapse or death, however, was only 7 months (2-20+). Major toxic effects included infections in 22 patients (grade 3 and 4 WHO), granulocytopenia (mainly grade 3) and nausea in 8 patients (mainly grade 1). 19/22 patients were in PR at the time of occurrence of infectious complications. Meanwhile, 14 patients died due to septicaemia, pneumonia or other infections. Nine patients developed severe septicaemia, 4 patients had pneumocystis carinii or aspergillus pneumonias. The high infection rate may not only be due to hypogammaglobulinaemia and granulocytopenia induced by FAMP but also to a remarkable decrease of CD4+ cells from a median of 2479 to 241 CD4+ cells/microliters after 6 cycles of FAMP. In one case a tumor lysis syndrome was observed. No CNS toxicity was noted. It is concluded that FAMP is effective even in patients with advanced CLL and low-grade NHLs refractory to multiple chemotherapy regimens. However, FAMP has a marked suppressive effect on granulocytes and T-lymphocytes, predominantly CD4+ lymphocytes.

Adult↗
PubMed1995

GM-CSF in a double-blind randomized, placebo controlled trial in therapy of adult patients with de novo acute myeloid leukemia (AML).

We investigated whether GM-CSF given concomitantly with chemotherapy (CT) and thereafter, improves the outcome of adults with de novo AML by increasing the efficacy of CT and reducing infections. CT included Ara-C, daunorubicin and etoposide (DAV) for induction and early consolidation therapy and one cycle with high-dose (patients aged < or = 50 years) or intermediate dose Ara-C (patients aged > 50 years)/daunorubicin for late consolidation therapy. Eight patients were randomized after DAVI to receive either GM-CSF (E. coli, 250 micrograms/m2/day, s.c.) or placebo starting 48 h prior to DAVII and the subsequent courses and given throughout CT until the absolute neutrophil count had recovered to > 500/microliters. The CR rate was 81% in the GM-CSF and 79% in the placebo group (p = 0.57; Fisher's exact test). The probability of relapse-free survival at 41 months after a median follow-up of 35 months was 42% in the GM-CSF and 41% in the placebo group (p = 0.89; log rank test). GM-CSF did not shorten the period of neutropenia < or = 500/microliters, while it prolonged the duration of thrombocytopenia < or = 25,000/microliters. The incidence and severity of infections as well as the non-hematological toxicity were similar in both groups. In summary, in this randomized trial GM-CSF as an adjunct to AML therapy was feasible. For the present GM-CSF does not have a significant effect on treatment outcome.

Adolescent↗
PubMed1995

The inhibition of lymphokine-activated killer cells in acute myeloblastic leukemia is mediated by transforming growth factor-beta 1.

In acute myeloblastic leukemia (AML), the T cell response and cytotoxic activity are impaired at time of diagnosis due to not-yet-identified soluble immunosuppressing factors. The inhibition of autologous antileukemic immune response by these factors may support immunosurveillance of AML. A well-known inhibitor of lymphokine-activated killer (LAK) cell activity is transforming growth factor-beta 1 (TGF-beta 1). To evaluate the possible significance of TGF-beta 1 for the impaired cytotoxic activity in AML at time of diagnosis, we looked for the TGF-beta 1-specific mRNA, for the production and release of TGF-beta 1, and for its relevance for immunosuppressing activities in AML. In the culture supernatants of 18 investigated AMLs, we detected various amounts of TGF-beta protein. The TGF-beta 1 and TGF-beta 2 protein concentrations were 105 pg/mL (< 50-240 pg/mL) and 32 pg/mL (< 2-91 pg/mL), respectively. In 13 of 15 patients, the leukemic blasts expressed TGF-beta 1 mRNA. To exclude possible interferences with contaminating mononuclear cells (MNC), the data were confirmed by analysis of sorted blast cells and leukemic cell lines. All investigated leukemic cell lines expressed TGF-beta 1 protein and mRNA. The culture supernatants of AMLs inhibited LAK activity strongly in a dose-dependent manner. The inhibition of cytotoxicity could be restored by the addition of neutralizing TGF-beta 1 antibodies. The data suggest TGF-beta 1 to be a relevant factor for the inhibition of cytotoxic activities in AMLs.

Adult↗
PubMed1995

Purging of peripheral blood stem cells yields BCR-ABL-negative autografts in patients with BCR-ABL-positive acute lymphoblastic leukemia.

Remission marrow from patients with BCR-ABL+ acute lymphoblastic leukemia (ALL) achieving clinical remission (CR) after induction or consolidation chemotherapy according to the German multicenter adult ALL (GMALL) protocol showed high titers of residual BCR-ABL+ cells. Therefore, we initiated a pilot study to monitor circulating BCR-ABL+ cells and to collect, purge, and autograft peripheral blood stem cells (PBSC) in these patients. After GMALL 05/93 high-risk phase II of induction chemotherapy (high-dose AraC 3 g/m2 x 8 does and mitoxantrone 10 mg/m2 x 3 doses), patients received 5-10 micrograms/kg subcutaneous recombinant human granulocyte colony-stimulating factor (rhG-CSF) daily. Mobilized CD34+ cells peaked between 20 and 26 days after starting chemotherapy at 4.8-75.6 (median 10.8) x 10(4)/mL peripheral blood (PB) (n = 5). Patients treated with additional chemotherapy cycles failed to mobilize adequate numbers of CD34+ cells. PB stem cells (PBSC) were purged using a cocktail of CD10, CD19, and AB4 monoclonal antibodies (mAbs) coupled to immunomagnetic beads (IMB). The median recoveries of total nucleated cells (TNC) and CD34+ cells after mAb/IMB purging were 84 and 81%. The peak numbers of CD34+ cells collected in a single leukapheresis were median 8.6 x 10(6)/kg pre- and 5.2 x 10(6)/kg postpurge (n = 4). The absolute prepurge CD19+ cells were as low as median 2.7 (range 1.4-19) x 10(6) per leukapheresis. Residual BCR-ABL+ cells in unpurged leukapheresis products were assessed by limiting-log10-dilution nested reverse-transcriptase polymerase chain reaction (RT-PCR) as one in 10(5) to one in 10(6) normal cells and were consistently undetectable in all purged PBSC autografts. We conclude that sufficient numbers of CD34+ cells for PBSCT can be collected after phase II but not at later stages of the GMALL 05/93 high risk protocol; PBSC grafts are 3 log less contaminated with residual BCR-ABL+ cells compared to an historical series of 13 autologous BM grafts; and purging of PBSC with mAb/IMB is feasible with minor loss of CD34+ cells and abolished BCR-ABL signals in the grafts.

Adult↗
PubMed1994

A low affinity chimeric human alpha/beta-granulocyte-macrophage colony-stimulating factor receptor induces ligand-dependent proliferation in a murine cell line.

The high affinity receptor for granulocyte-macrophage colony-stimulating factor (GM-CSF) is composed of at least two subunits, an 85-kDa low affinity GM-CSF-binding protein (alpha-GMR) and a 120-kDa beta-subunit (beta-GMR) necessary for high affinity binding and signal transduction. Previous studies have shown that deletion of the intracellular domain of alpha-GMR inactivates the receptor's ability to support proliferation, but has no effect on GM-CSF binding. Using anti-alpha-GMR- and anti-beta-GMR-specific antibodies, we show that alpha-GMR and beta-GMR coprecipitate only after GM-CSF binding, suggesting that binding of GM-CSF induces stabilization or assembly of an activated receptor complex involving recruitment of beta-GMR chains. To understand the contribution of each subunit of this receptor to the generation of an activated receptor complex, we attempted to construct minimal receptors with some or all of the functions of the wild-type heterodimer. We found that a hybrid human alpha/beta-GMR molecule in which the extracellular and transmembrane segments are composed of alpha-GMR sequences and the intracellular segment is composed of beta-GMR bound GM-CSF with low affinity, but activated tyrosine kinase activity, induced receptor internalization, and supported short- and long-term proliferation of transfected Ba/F3 cells. At least 1 ng/ml human GM-CSF was required for growth stimulation, and maximal proliferation occurred at a concentration of 10 ng/ml. This was 10-100-fold more than needed to stimulate growth of Ba/F3 cells expressing both full-length human alpha-GMR and beta-GMR and 1000-fold less than needed to stimulate growth of Ba/F3 cells expressing only human alpha-GMR. These results indicate that the cytoplasmic domain of alpha-GMR is not required to initiate a unique signaling event for proliferation in Ba/F3 cells, but can be functionally replaced by the cytoplasmic domain of beta-GMR.

3T3 Cells↗