The current and future role of the medical oncologist in the professional care for cancer patients: a position paper by the European Society for Medical Oncology (ESMO).

The number of cancer patients in Europe is rising and significant advances in basic and applied cancer research are making the provision of optimal care more challenging. The concept of cancer as a systemic, highly heterogeneous and complex disease has increased the awareness that quality cancer care should be provided by a multidisciplinary team (MDT) of highly qualified healthcare professionals. Cancer patients also have the right to benefit from medical progress by receiving optimal treatment from adequately trained and highly skilled medical professionals. Built on the highest standards of professional training and continuing medical education, medical oncology is recognised as an independent medical specialty in many European countries. Medical oncology is a core member of the MDT and offers cancer patients a comprehensive and systemic approach to treatment and care, while ensuring evidence-based, safe and cost-effective use of cancer drugs and preserving the quality of life of cancer patients through the entire 'cancer journey'. Medical oncologists are also engaged in clinical and translational research to promote innovation and new therapies and they contribute to cancer diagnosis, prevention and research, making a difference for patients in a dynamic, stimulating professional environment. Medical oncologists play an important role in shaping the future of healthcare through innovation and are also actively involved at the political level to ensure a maximum contribution of the profession to Society and to tackle future challenges. This position paper summarises the multifarious and vital contributions of medical oncology and medical oncologists to today's and tomorrow's professional cancer care.

[Venous thrombembolism in tumour patients]. / Venöse Thrombembolien bei Tumorpatienten.

Venous thrombembolism (VTE) is one of the most frequent complication in cancer patients. The current options in prophylaxis and therapy have to be balanced against the risks of major bleeding and the burden for the patients. The Gesellschaft für Thrombose- und Hämostaseforschung, the Deutsche Gesellschaft für Palliativmedizin and the German speaking Societies of Hematology and Oncology have recently published guidelines on VTE in cancer patients. Recommendations include diagnostics, individual prophylaxis and treatment.

Role and responsibility of oncologists in assisted suicide. Practice and views among members of the German Society of Haematology and Medical Oncology.

BACKGROUND: Physician-assisted suicide (PAS) is a controversial practice and regulatory frameworks differ regarding assigned physicians' roles. This study explores clinical experience and views of German oncologists concerning ethically and legally relevant aspects of PAS after change of the law. MATERIALS AND METHODS: An online survey was conducted among members of the German Society of Haematology and Medical Oncology (DGHO) in March 2021. Descriptive analysis, bivariate and multivariable logistic regression of quantitative data on determinants related to (un)willingness to assist with suicide as well qualitative analysis of free-text comments were carried out. RESULTS: Seven hundred and forty-five of 3588 DGHO members responded (20.8%). Of these, 29.9% reported requests for a lethal drug and 3.0% (n = 22) reported to have assisted with suicide. Almost half of them (47.0%, n = 350) objected to providing PAS, whereas 45.9% indicated a willingness at least under certain conditions. Of those respondents who did not object to PAS, 25.4% would also consider assistance if those willing to die had a psychiatric disease and 10.2% if requestors had no disease at all. A majority viewed a role for physicians regarding different tasks associated with assisted suicide. Respondents with <10 years of professional experience, working in hospital with religious affiliation and with subspecialisation in palliative care were significantly less frequently willing to assist suicide. CONCLUSIONS: Respondents are divided in their personal attitudes towards PAS but a majority supports involvement of physicians regarding different tasks related to assisted suicide. Data about the practice and envisaged professional role may inform development of an acceptable ethico-legal framework for a controversial practice.

A four-gene LincRNA expression signature predicts risk in multiple cohorts of acute myeloid leukemia patients.

Prognostic gene expression signatures have been proposed as clinical tools to clarify therapeutic options in acute myeloid leukemia (AML). However, these signatures rely on measuring large numbers of genes and often perform poorly when applied to independent cohorts or those with older patients. Long intergenic non-coding RNAs (lincRNAs) are emerging as important regulators of cell identity and oncogenesis, but knowledge of their utility as prognostic markers in AML is limited. Here we analyze transcriptomic data from multiple cohorts of clinically annotated AML patients and report that (i) microarrays designed for coding gene expression can be repurposed to yield robust lincRNA expression data, (ii) some lincRNA genes are located in close proximity to hematopoietic coding genes and show strong expression correlations in AML, (iii) lincRNA gene expression patterns distinguish cytogenetic and molecular subtypes of AML, (iv) lincRNA signatures composed of three or four genes are independent predictors of clinical outcome and further dichotomize survival in European Leukemia Net (ELN) risk groups and (v) an analytical tool based on logistic regression analysis of quantitative PCR measurement of four lincRNA genes (LINC4) can be used to determine risk in AML.

Increasing intensity of therapies assigned at diagnosis does not improve survival of adults with acute myeloid leukemia.

We randomized 3375 adults with newly diagnosed acute myeloid leukemia (AML) or high-risk myelodysplastic syndrome to test whether increasingly intensive chemotherapies assigned at study-entry and analyzed on an intent-to-treat basis improved outcomes. In total, 1529 subjects <60 years were randomized to receive: (1) a first course of induction therapy with high-dose cytarabine and mitoxantrone (HAM) or with standard-dose cytarabine, daunorubicin and 6-thioguanine (TAD) followed by a second course of HAM; (2) granulocyte-colony stimulating factor (G-CSF) or no G-CSF before induction and consolidation courses; and (3) high-dose therapy and an autotransplant or maintenance chemotherapy. In total, 1846 subjects ⩾60 years were randomized to receive: (1) a first induction course of HAM or TAD and second induction course of HAM (if they had bone marrow blasts ⩾5% after the first course); and (2) G-CSF or no G-CSF as above. Median follow-up was 7.4 years (range, 1 day to 14.7 years). Five-year event-free survivals (EFSs) for subjects receiving a first induction course of HAM vs TAD were 17% (95% confidence interval, 15, 18%) vs 16% (95% confidence interval 14, 18%; P=0.719). Five-year EFSs for subjects randomized to receive or not receive G-CSF were 19% (95% confidence interval 16, 21%) vs 16% (95% confidence interval 14, 19%; P=0.266). Five-year relapse-free survivals (RFSs) for subjects <60 years receiving an autotransplant vs maintenance therapy were 43% (95% confidence interval 40, 47%) vs 40 (95% confidence interval 35, 44%; P=0.535). Many subjects never achieved pre-specified landmarks and consequently did not receive their assigned therapies. These data indicate the limited impact of more intensive therapies on outcomes of adults with AML. Moreover, none of the more intensive therapies we tested improved 5-year EFS, RFS or any other outcomes.

Management of acute myeloid leukemia in elderly patients.

Acute myeloid leukemia (AML) at older age is associated with several biologic and clinical characteristics. Hence, it may arise from an early level of hematopoietic stem cells and has a high frequency of blast cells with multidrug resistance glycoprotein MDR1 expression and particularly a high incidence of poor prognostic karyotypes. These factors, rather than age per se, underlie the poorer outcome as compared with younger cases. Prospective randomized studies clearly demonstrate, however, that elderly patients benefit from more intensive induction therapy and particularly from full-dose application of anthracyclines and possibly also cytarabine. Hematopoietic growth factors accelerate the recovery from treatment-induced neutropenia and may improve the remission rate, remission duration, and even overall survival. New treatment strategies need to be developed, however, for poor-prognosis AML subtypes in order to further improve the therapeutic perspectives for elderly patients with AML.

Towards a pathogenesis-oriented therapy of acute myeloid leukemia.

Genetic and molecular techniques have provided increasing insights into the biology of acute myeloid leukemia (AML). These investigations showed that AML is not a homogeneous disease but a heterogeneous group of biologically different subentities. These subentities are currently primarily defined by cytogenetics by which three main subgroups can be discriminated: AML with balanced translocations, AML with unbalanced aberrations and AML without cytogenetically detectable aberrations. Within the latter group molecular alterations are identified in more than half of cases such as NPM mutations, FLT3 mutations, MLL duplications and mutations of CEBP-alpha. The clinical meaning of these findings is illustrated by substantial differences in response to therapy and long-term outcome. As demonstrated by the recent multicenter trial of the German AML Cooperative Group (AMLCG) and other studies intensification of induction therapy may improve the results in distinct subtypes but fails to do so in others. Therefore, new strategies need to be explored which incorporate the knowledge about the biology of AML to develop biology adapted treatment strategies. This process has just begun and is predominantly determined by the availability of new agents and their evaluation in clinical phase I and II studies. A variety of targets are currently explored and some trials have yielded promising results already. The step towards a biology adapted treatment of AML is long and requires the combined efforts of researchers, clinicians and the pharmaceutical industry. The first steps towards this goal have been taken and give rise to the hope for more effective and more specific therapies of AML.

Analysis of expression and function of CD40 on normal and leukemic human B cell precursors.

CD40 was originally identified on circulating and tonsillar human B cells, and anti-CD40 monoclonal antibodies (MoAb) are known to deliver a progression signal to activated B cells. However, the expression and function of CD40 on human B cell precursors (BCP) have not been examined in detail. Two new anti-CD40 MoAb were produced and shown to recognize an epitope indistinguishable from the anti-CD40 antibody G28-5. CD40 was readily detected on normal and leukemic BCP by flow cytometry, and cell surface expression was upregulated by phorbol ester. Despite the ability of normal and leukemic BCP to respond to phorbol ester (PMA) and/or low molecular weight B cell growth factor (L-BCGF), anti-CD40 exerted no stimulatory action and could not enhance the response of these cells to PMA, L-BCGF, or both. Cross-linking anti-CD40 MoAb with rabbit anti-mouse Ig also failed to induce a proliferative response in normal BCP. We conclude that anti-CD40 does not exert demonstrable agonistic effects on normal and leukemic human BCP. Our results suggest that signal delivery through CD40 and/or subsequent intracellular signal processing may require accessory molecules not expressed in BCP, or CD40 may subserve a different function for BCP.

Structure/function analyses of IL-2 binding proteins on human B cell precursor acute lymphoblastic leukemias.

We have studied the expression and function of interleukin-2 (IL-2) receptors on B cell precursor acute lymphoblastic leukemias (ALL). After incubation of B cell precursor ALL in vitro for 24 hr, 11 out of 17 leukemic bone marrow aspirates expressed the Tac/CD25 protein (greater than 10% positive blasts). Expression of Tac/CD25 on the leukemic cells was confirmed by two color flow cytometric analysis using anti-Tac/CD25 and anti-CALLA/CD10 monoclonal antibodies. The molecular mass of the B cell precursor ALL Tac/CD25 protein was 55 kilodaltons (kD), identical to that on activated T cells. Binding of radiolabeled IL-2 in two leukemic bone marrow aspirates demonstrated the presence of high affinity IL-2 receptors. Cross-linking of 125I-labeled IL-2 to TPA activated B cell precursor ALL revealed the 55 kD Tac/CD25 protein and an additional protein of 75 kD. Recombinant IL-2 in concentrations of 10-1,000 U/ml had essentially no proliferative effect in 10 patients tested, whereas low molecular weight B cell growth factor (L-BCGF) induced proliferation in 8 of 10 patients. L-BCGF also induced expression of CD20 in 3 of 7 CD20 negative B cell precursor ALL. IL-2 did not induce CD20, but enhanced its expression in the 3 patients who responded to L-BCGF. We conclude that IL-2 has essentially no proliferative effect on B cell precursor ALL, despite the presence of high affinity IL-2 receptors and the presence of the IL-2 binding cell surface molecules similar to those on activated T cells. IL-2 may, however, induce a phenotypic change (CD20 acquisition) consonant with differentiation in synergy with L-BCGF.

Clonal chromosomal abnormalities in the stem cell compartment of patients with acute myeloid leukemia in morphological complete remission.

Acute myeloid leukemia arises from the clonal expansion of a malignant transformed progenitor cell. Despite intensive chemotherapy, final disease eradication is achieved by a small proportion of cases only and 50-70% of adults with AML will ultimately relapse and die from their disease. Hence residual disease below the level of morphological detectability must be assumed in clinical and morphological complete remission. CD34+/CD38- and CD34+/CD38+ subpopulations of seven patients in morphological complete remission were isolated by FACS (purity >98%) and were analyzed by conventional cytogenetics or FISH for chromosomal aberrations. In five of seven patients, clonal chromosomal abnormalities were detected in the CD34+/CD38+ subpopulation and in one patient with AML M2 (add (2)(q37)) in the most immature CD34+/CD38- stem cell compartment. One patient with AML M4Eo (inv(16),+8), showed a normal karyotype by conventional cytogenetic analysis, whereas four of 15 metaphases of the sorted CD34+/CD38+ subpopulation revealed the inversion 16. These observations underline that leukemic cells can survive intensive chemotherapy in the niche of the stem cell compartment. In some patients the sensitivity for the detection of persistent leukemic cells seems to be higher in FACS-sorted subpopulations than conventional cytogenetic analysis of the unseparated bone marrow. Immunophenotyping revealed minimal residual disease in four of the patients. Functional analysis has to be performed to investigate the leukemogenic potential of these residual cells.

Proliferative effect of interleukin-3 on normal and leukemic human B cell precursors.

We have examined the influence of recombinant, human IL-3 on normal and leukemic B cell precursors. CD10/CALLA+ leukemic B cell precursors, the pre-B cell line BLIN-1, and surface IgM-B cell precursors isolated from fetal bone marrow all responded to IL-3. This IL-3 response was dose-dependent and could be abrogated by a rabbit anti-IL-3 antiserum. Binding studies using radiolabeled IL-3 revealed the presence of approximately 200 high affinity IL-3 receptors on the BLIN-1 cell line, with a KD of 150pM. We conclude that normal and leukemic human B cell precursors express functional IL-3 receptors, extending the biologic range of this hematopoietin to the lymphoid lineage.

Differential effect of GM-CSF pretreatment on intracellular Ara-C metabolism in normal bone marrow mononuclear cells vs acute myeloid leukemia (AML) blasts.

The effect of recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF) on the intracellular metabolism of cytosine arabinoside (Ara-C) was comparatively analyzed in normal bone marrow mononuclear cells (NBMMC) from eight healthy volunteers and in leukemic blasts from 50 patients with acute myeloid leukemia (AML). Pretreatment with GM-CSF (100 U/ml) for 48 h resulted in a significant enhancement of DNA synthesis in both cell types: 21 of 35 AML specimens were found to be responsive to GM-CSF as defined by an increase of 3H-TdR incorporation into the DNA > 1.5-fold while NBMMC from normal donors were responsive in all cases. In GM-CSF responsive AML blasts, overall DNA polymerase and DNA polymerase alpha activity increased from a median of 84.4 to 96.1 and from 3.45 to 5.2 pmol/min x mg as compared to a median of 96.7 to 189.9 and 1.2 to 2.2 pmol/min x mg in NBMMC (P < 0.05). Median Ara-C-mediated inhibition of DNA synthesis was significantly more effective in AML blasts as compared to NBMMC (76.5 vs 55.0% at 0.05 microM and 99.0 vs 96.0% at 5.0 microM Ara-C, P < 0.01) but was not influenced by GM-CSF pretreatment. Similarly, intracellular Ara-CTP levels were higher in AML blasts as compared to NBMMC (median of 46.9 vs 18.7 at 1 microM, 167.8 vs 48.0 at 10 microM and 337.5 vs 59.5 ng/10(7) cells at 100 microM extracellular Ara-C, P < 0.01) but showed no enhancement in the presence of GM-CSF. Median deoxycytidine (DCK) and thymidine kinase (TK) activity were only slightly increased in AML blasts after GM-CSF priming. In contrast, NBMMC revealed a significant increase in TK activity after GM-CSF pretreatment (from a median of 1.9 to 3.6 pmol/min x mg, P = 0.039). At low; intermediate and high extracellular Ara-C concentrations GM-CSF pretreatment resulted in a significant enhancement of the 3H-Ara-C incorporation into the DNA in both GM-CSF responsive AML blasts and NBMMC (median of 1.3 to 2.1- and 1.4 to 1.6-fold, P < 0.05). GM-CSF non-responsive AML blasts showed no change in 3H-Ara-C incorporation into the DNA in response to GM-CSF at low Ara-C concentrations but significant increases at intermediate and high extracellular Ara-C concentrations (median increases of 1.63-fold at 1.06 microM with P = 0.01 and 1.37-fold at 10 microM extracellular Ara-C with P = 0.0+005). NBMMC revealed significantly lower GM-CSF-induced increases of the 3H-Ara-C incorporation into the DNA as compared to the effect of GM-CSF priming on DNA synthesis (median increases of 1.4 to 1.7-fold vs 2.6-fold, P < 0.05). These data reveal a different effect of GM-CSF priming on the metabolism of Ara-C in normal vs leukemic cells which may cause a preferential increase in the antileukemic cytotoxicity of Ara-C in the presence of GM-CSF.

Blast cell proliferative activity and sensitivity to GM-CSF in vitro are associated with early response to TAD-9 induction therapy in acute myeloid leukemia.

The current study was undertaken to determine the relevance of leukemic blast cell proliferative activity, cellular parameters of Ara-C metabolism and the in vitro sensitivity to GM-CSF in association with the clinical response to TAD-9 induction therapy in 66 patients with de novo acute myeloid leukemia (AML). Proliferative activity was assessed by 3H-thymidine (3H-TdR) incorporation and thymidine kinase (TK) activity, parameters of Ara-C metabolism comprised the activities of deoxycytidine kinase (DCK) and DNA polymerase alpha (poly alpha) as well as Ara-CTP concentrations and 3H-Ara-C uptake into DNA. GM-CSF sensitivity was determined by in vitro incubation of blasts for 48 h with or without GM-CSF (100 U/ml) followed by an additional 4 h concurrent exposure to GM-CSF and 3H-TdR (0.5 microCi/ml). The following results were obtained as expressed by median values and ranges: 3H-TdR incorporation: 1.07 pmol/10(5) cells (0.0-10.1), TK: 7.3 pmol/min/mg protein (1.3-56.0), DCK: 9.3 pmol/min/mg protein (0.77-47.1), poly alpha: 1.7 pmol/min/mg protein (0.00-28.9), Ara-CTP: 53.3 ng/10(7) cells (13.3-211.0), 3H-Ara-C uptake: 0.06 pmol/10(5) cells (0.0-0.57). 3H-Ara-C uptake was correlated with 3H-TdR incorporation (r = 0.74) and with the (S-phase dependent) activities of TK (r = 0.73) and poly alpha (r = 0.71, but not with DCK activity or intracellular Ara-CTP content. Blast cells of 37 from 55 analyzed patients were found to be sensitive to GM-CSF stimulation as defined by an increase in 3H-TdR incorporation > or = 1.5-fold over control values after the 48 h GM-CSF exposure. In vitro data were related with clinical response to TAD-9 induction therapy in 43 patients with newly diagnosed AML, taking the blast cell reduction at day 10 or 16 to < 5% or > or = 5% residual blasts as early parameter for adequate or inadequate response, respectively. While neither 3H-Ara-C uptake, nor intracellular Ara-CTP concentration, TK nor DCK activity were predictive for response, a high 3H-TdR incorporation and a high poly alpha activity were associated with adequate blast cell reduction. Median values of 3H-TdR incorporation were 2.26 pmol/10(5) cells for patients with adequate blast cell clearance and 0.80 pmol/10(5) cells for patients with inadequate blast cell clearance (P = 0.11), the respective values for poly alpha were 3.22 pmol/min/mg protein for responders and 1.1 pmol/min/mg protein for non-responders (P = 0.0085).(ABSTRACT TRUNCATED AT 400 WORDS)

Trisomy 4 in 'stem cell-like' leukemic cells of a patient with AML.

CD34 positive progenitor cells were analyzed in the bone marrow aspirate from a patient with newly diagnosed AML FAB M4. The patient had trisomy 4 as sole cytogenetic abnormality and a dominant population of CD34 negative leukemic blasts. Karyotyping of the FACS isolated, minor subpopulation of CD34+/CD38-, 'stem cell'-like cells (incidence 0.29%) revealed trisomy 4 in 11/13 metaphases. No metaphases were obtained in the CD34 negative subpopulation. The experiments point to the existence of leukemic stem cells in the CD34+/CD38- compartment in AML patients with trisomy 4.

Multivariate analysis of prognostic factors in patients with refractory and relapsed acute myeloid leukemia undergoing sequential high-dose cytosine arabinoside and mitoxantrone (S-HAM) salvage therapy: relevance of cytogenetic abnormalities.

To improve the basis for the stratification of patients with refractory and relapsed acute myeloid leukemia (AML) univariate and multivariate analyses of prognostic factors were performed in 254 patients (median age 50 years, range 18-74) undergoing S-HAM salvage chemotherapy during two consecutive prospective trials of the German AML Cooperative Group. In a multivariate analysis, duration of the first complete remission (CR) was the only factor associated with time to treatment failure (P = 0.0223). Disease-free survival was influenced by a short duration of the first CR of less than 6 months (P = 0.0001), WBC (P = 0.0018), blast count (P = 0.0037), and neutrophil count (P = 0.0119). The achievement of CR was related to the hemoglobin level only (P = 0.0457), the early death rate was related to age only (P = 0.0109), and survival was related to the bilirubin level only (P = 0.0166). In the subgroup of 104 patients in whom additional karyotype analyses were performed prior to first-line therapy unfavorable chromosome abnormalities were associated with a lower CR rate (univariate analysis, P = 0.0342; CR 24% vs 53%) and were the only factor related to survival. These analyses warrant the further evaluation of the impact of cytogenetic abnormalities on the outcome of patients with advanced AML in order to improve the characterization according to duration of first CR and to WBC of distinct subgroups of patients with differing prognoses as a basis for stratification in future trials.

Cytogenetic analysis of CD34+ subpopulations in AML and MDS characterized by the expression of CD38 and CD117.

It has been supposed in de novo AML that malignant transformation occurs at the level of committed progenitors. Recent data of our group and others provide evidence that in AML malignant transformation may regularly occur at the level of stem cells. These cells can be discriminated by function and specific surface molecules. CD34, a glycophosphoprotein, is a cellular surface antigen characteristically expressed by stem cells. CD34+ stem cells can be further subdivided by the expression of additional surface molecules like CD38 and CD117. In this article we present results from cytogenetic examinations of FACS-isolated stem cell subpopulations in eight patients (four AML and four MDS). Six of them displayed clonal karyotype abnormalities at the time of first diagnoses in the native bone marrow (5q-; 5q- and complex abnormalities; +8; inv(16) and +8; i(17q) and -21; i(21q)). We used CD117, the receptor for the stem cell factor (also KIT oncogene) as a new cellular surface marker. CD34+/CD117+/- stem cell subpopulations were examined in two patients with AML and three patients with MDS. We found leukemic stem cells in every type of stem cell subpopulation examined (CD34+/CD38-, CD34+/CD38+, CD34+/CD117-, CD34+/CD117+). Secondary, progression-associated chromosome abnormalities likewise were demonstrable in CD34+ cells. In three patients a mosaic of normal and abnormal metaphases was found in the highly purified stem cell subpopulations. We conclude that in AML and MDS stem cells are the target of leukemogenic genetic defects. CD117 as a new marker to isolate different CD34+ subpopulations was not sufficient to discriminate between normal and leukemic stem cells. Our findings have implications for autologous stem cell transplantation, high-dose chemotherapy and the pathogenetic concept of leukemogenesis.

In vitro activation of low-grade non-Hodgkin's lymphoma by murine fibroblasts, IL-4, anti-CD40 antibodies and the soluble CD40 ligand.

The in vitro analysis of growth regulation in low-grade B non-Hodgkin's lymphoma (B-NHL) is hampered by the rapid apoptotic death of the malignant B cells ex vivo. A complex culture system, using murine CDw32 transfected fibroblasts (LTK-cells), IL-4 and anti-CD40 mAb, has been established for the propagation of normal mature B cells in vitro. We investigated the influence of the different components of this coculture system on cell survival and apoptosis of B-NHL cells. Nine samples from patients with follicular lymphoma and from eight patients with immunocytoma were analyzed. No cell proliferation of B-NHL cells could be induced in the culture system. However, CDw32-transfected murine fibroblasts most efficiently supported cell viability of B-NHL cells with an increase in cell survival by 114% compared to the control (P = 0.047). IL-4 alone also had a stimulatory effect on cell survival of B-NHL cells after 6 days. In contrast, the soluble recombinant CD40 ligand gp39 and the anti-CD40 mAbs mAb89 and EA-5 did not prolong cell survival. CDw32 transfectants blocked apoptosis of B-NHL cells efficiently from 67% in the control to 16% (P = 0.001). Reduction in apoptosis was accompanied by an elevated bcl-2 protein expression. IL-4 or mAb89 did not further reduce apoptotic cell death in CDw32 transfectant-dependent cocultures. Our data underline the pivotal role of LTK- cells for cell survival of B-NHL cells in vitro. The efficient blockage of apoptosis associated with increased bcl-2 protein expression causes prolonged cell viability of the B-NHL cells.

TGF-beta inhibits growth and induces apoptosis in leukemic B cell precursors.

The uncontrolled proliferation of malignant lymphoblasts is the pathobiological hallmark in B cell precursor-ALL (BCP-ALL). Identification of inhibitory growth factors is of great importance for the understanding of growth control of leukemic B cell precursors and the development of novel therapeutic approaches in BCP-ALL. The aim of our study was the analysis of the effect of TGF-beta on cell survival and apoptosis of B cell precursors (BCP) from patients with acute lymphoblastic leukemia in vitro. Experiments were performed in a coculture system with cloned murine fibroblasts, which efficiently block spontaneous ex vivo apoptosis of BCP and thus allows the assessment of cytokine-induced growth inhibition. TGF-beta significantly reduced cell viability of highly purified, FACS isolated CD10+/CD19+ leukemic BCP by a mean of 53% (P = 0.0001). The loss of cell viability was accompanied by a significant increase of apoptosis with a mean of 70% (P = 0.0028). The TGF-beta effect was blocked specifically by a monoclonal anti-TGF-beta antibody. Induction of apoptotic cell death by TGF-beta was not accompanied by reduction of bcl-2 protein expression. TGF-beta transcription was not detected in the leukemic pre-B cell line BLIN-1, but in the murine fibroblasts. The growth inhibitory effect of TGF-beta was not restricted to leukemic BCP. The cytokine also increased apoptosis of normal, highly purified BCP by a mean of 58%. The data identify TGF-beta as a potent growth inhibitory cytokine for leukemic BCP.

Longterm effects of prolonged maintenance and of very early intensification chemotherapy in AML: data from AMLCG.

In order to further improve the cure rate in AML we investigated the effect of more chemotherapy--in terms of its intensity and its duration--in 2 studies. In our 1981 study patients received TAD 1-2 courses for induction, 1 course for consolidation and randomly no further treatment or monthly myelosuppressive maintenance for 3 years. Evaluating 213 responders remission duration was clearly longer in the maintenance group with 24% CCR after 5 and 10 years. In our 1985 study the same successful strategy was further intensified by a second induction course given regardless of response to the first course to all patients up to 60 years of age while older patients received standard induction as before. This age-adapted concept resulted in a further increase of 5 years CCR in the 461 responders to as much as 34% not achieved for unselected patients in other multicenter trials. 20 patients receiving auto-BMT in first CR show the same relapse free survival as their counterparts receiving chemotherapy according to the 1985 protocol in a matched-pair analysis. We conclude that both very early intensification and prolonged maintenance contribute to a higher cure rate that is not further improved even by a maximum intensity short-term treatment. The limits of chemotherapy in AML may be overcome by modulating its myelotoxicity and antileukemic potency using GM-CSF as shown in 2 studies of our group.

Flow cytometric characterization of acute myeloid leukemia. Part II. Phenotypic heterogeneity at diagnosis.

The frequency and distribution of aberrant antigen expression are analyzed on bone marrow aspirates from 80 patients with newly diagnosed acute myeloid leukemia (AML) by multidimensional flow cytometry. Parameters examined are the light scatter profile of the leukemic cells and the correlative expression of different combinations of the CD2, 4, 5, 7, 11b, 11c, 13, 14, 15, 16, 33, 34, 38, and HLA-DR antigens. Antigen expression on leukemic cells in bone marrow is described by characteristic antigen expression patterns describing: (i) the percentage of cells expressing the antigen; (ii) the antigen density; and (iii) the distribution of the antigen on the leukemic cells. Typically the non-myeloid antigens are homogeneously expressed by the leukemic cells, whereas the myeloid associated antigen CD11b, CD11c, CD14, and CD15 are heterogeneously expressed. Comparison of the antigenic profiles of 80 bone marrow aspirates revealed an extreme interclonal heterogeneity. Comparison of the antigen expression patterns found in AML patients with the antigen expression in normal bone marrow revealed four patterns of aberrant antigen expression in AML: (i) expression of nonmyeloid antigens (i.e. CD2, CD5, and CD7 were present in 57, 60, and 37% of the patients, respectively); (ii) asynchronous expression of myeloid associated antigens (i.e. co-expression of CD34 and CD15 in 25% of the patients and expression of CD16 on immature myeloid cells in 15% of the cases); (iii) over-expression of myeloid associated antigens (e.g. CD34 in 16% of the cases and CD14 on neutrophilic cells in 19% of all patients); and (iv) absence of expression of myeloid associated antigens (e.g. lack of CD33 in 21% of the cases and lack of both CD11b and CD15 in 6% of all patients. Multidimensional flow cytometric analysis of bone marrow aspirates of AML patients disclosed that the leukemic cells of each AML patient had a unique antigenic profile and could be discriminated from their normal counterparts based on aberrant antigen expression and typical light scatter profiles. The ability to distinguish leukemic cells from normal cells allows the detection of residual leukemic cells during and after chemotherapy.