Otto Kloke

Lupus-like autoimmune disease induced by interferon therapy for myeloproliferative disorders

Symptoms of autoimmune disease were evaluated in 125 patients with chronic myelogenous leukemia (CML) and in 12 patients with essential thrombocythemia undergoing treatment with recombinant interferon (IFN)-alpha-2b plus/minus low-dose recombinant IFN-gamma. Twenty-seven of 137 patients (20%) developed rheumatoid symptoms. Furthermore, the incidence of antinuclear antibody (ANA) formation was studied. Elevated ANA titers were found in 5/19 (26%) of CML patients at the time of diagnosis and in 3/18 (17%) of patients treated with hydroxyurea or busulfan. During IFN treatment, 18 of 25 tested patients (72%) had elevated ANA titers. In 15 of these ANA-positive patients, clinical signs of autoimmune disease appeared. All these patients were under long-term IFN treatment and were in remission of disease. In three patients criteria for systemic lupus erythematosus were fulfilled. Severity of side effects had led to the discontinuation of IFN treatment in these patients. The data indicate that IFN-alpha and IFN-gamma can induce ANA associated with autoimmune disease in patients with myeloproliferative disorders.

Impact of interferon alpha-induced cytogenetic improvement on survival in chronic myelogenous leukaemia

Summary. The objective of this study was to investigate the prognostic impact of the reduction of Philadelphia chromosome (Ph) positive metaphases by treatment of chronic myelogenous leukaemia (CML) with interferon (IFN) alpha. Therefore, we evaluated the outcome of patients with previously untreated chronic phase Ph‐positive CML, enrolled from 1984 to 1990 into two consecutive IFN trials at our institution. Of a total of 71 patients, 62 (87%) were evaluable for cytogenetic response. No cytogenetic improvement was seen in 16 patients (23%), 28 patients (38%) had a decrease in Ph‐positive bone marrow metaphases to levels ranging from 35% to 95%, and nine patients (13%) to levels between 5% and 34%. In nine patients (13%), Ph‐positive metaphases were no longer detectable. After a median follow‐up period of 33 months, the projected 5‐year survival is 55% for the 62 patients evaluable for cytogenetic response. In this patient population there was no significant difference in the survival probability according to patients’risk status as defined by the Sokal score. Categorization according to the extent of Ph reduction, however, allowed three groups with significantly different prognoses to be identified. Patients achieving a Ph reduction to less than 35% were found to constitute a low risk group with a median survival not yet known and a projected 5‐year survival of 90%. The 5‐year survival rate was 55% for patients with a Ph reduction to levels between 35% and 95%, and less than 10% for those without any cytogenetic improvement. Thus, this study demonstrates that cytogenetic improvement on IFN treatment is an important prognostic factor for survival.

Combination therapy with interferon alpha‐2b plus low‐dose interferon gamma in pretreated patients with Ph‐positive chronic myelogenous leukaemia

Between March 1988 and July 1990, 28 adults with chronic myelogenous leukaemia (CML) were treated with a combination of recombinant human interferon (IFN) alpha‐2b s.c. (initial dose 4 x 106 U/m2) and recombinant human IFN gamma s.c. (50 μg totally) daily. All patients were in chronic phase disease and had been treated previously with chemotherapy or bone marrow transplantation. A complete haematologic remission was achieved in three patients (11%), a haematologic remission in 12 patients (43%), and a partial haematologic remission in seven patients (25%). Six patients did not respond to this schedule. Acute side‐effects were flu‐like symptoms, fever and chills. During long‐term treatment six patients developed poly‐arthralgia. Haematotoxicity WHO grade III occurred in three patients, and WHO grade IV in two patients. One patient developed psychosis, and in another patient an exacerbation of a pre‐existing sarcoidosis was observed. We conclude that this combination is tolerable and effective in inducing haematological remissions in pretreated CML patients.

Interferon alfa-2b in acute- and chronic-phase chronic myelogenous leukaemia: Initial response and long-term results in 54 patients

Fifty-four patients with Ph1-positive chronic myelogenous leukaemia (CML) (48 with chronic-phase and six acute-phase disease) were treated with interferon alfa-2b subcutaneously (s.c.). The starting dose was 4 million units (MU)/m2 body surface area daily. It was reduced in parallel with serially determined leucocyte counts, and minimal effective doses were given as maintenance after achieving remission. Haematological remissions were induced in 22 of the 48 patients (46%) with chronic-phase disease. Thirteen patients (27%) revealed partial haematological remission and another 13 no response to treatment. No complete remission could be induced, although minor or partial cytogenetic responses were seen in 16 patients (33%). Moreover, a bcr-abl reduction was detected on Southern blot analysis in two patients. In chronic-phase disease, results of treatment were influenced by elapsed time after diagnosis, extent of previous treatment and interferon dosage. No beneficial effects of interferon were detected in the six patients with acute-phase disease. Principal acute side effects were fever and flu-like symptoms at the beginning of the therapy, which usually subsided within 3-7 days. Chronic side effects, especially weakness and neuropathy, were less frequent but more severe and necessitated discontinuation of treatment in 10 patients. In summary, interferon alfa-2b seems to be an effective treatment in early chronic-phase CML. Long-term effects on the course of the disease, however, must be determined.

Frequency of Clonal B Lymphocytes in Chronic Myelogenous Leukemia Evaluated by Fluorescence In Situ Hybridization

The demonstration of the Philadelphia (Ph) chromosome in B lymphocytes from patients with chronic myelogenous leukemia (CML) has provided evidence that the disorder originates in a pluripotent progenitor cell. Divergent results, however, exist as to the degree of contribution of clonally derived cells to the B-cell compartment. To address this issue, B lymphocytes were selected from the blood of seven patients in the chronic phase of Ph-positive CML and were examined with dual-color fluoresence in situ hybridization for the presence of the Ph translocation. The purity of the B-cell preparations ranged from 88% to 97% (mean 93%). The Ph translocation was detected in 22-34% (mean, 27%) of the sorted B cells. There was no evidence that the duration of the disease affects the ratio of Ph-positive and -negative B cells. In summary, clonally derived circulating B lymphocytes were present in all patients studied but made only minor contribution to this compartment.

Tumor necrosis factor α modifies resistance to interferon α in vivo: First clinical data

SummaryPatients with Philadelphia-positive chronic-phase chronic myelogenous leukemia (CML) resistant to interferon (IFN) α were treated in a phase I/II study with recombinant human tumor necrosis factor α to overcome IFNα resistance. Doses of 40, 80, 120 or 160 µg/m2 TNFα were given as 2-h infusions on 5 consecutive days every 3 weeks. IFNα (4 × 106 IU/m2 s.c., daily) treatment was continued. Six patients were treated, completing 1–24 (median, 12) treatment cycles. Five of the six patients achieved partial hematological remission, while the remaining patient had to stop treatment because of WHO grade 4 thrombocytopenia following the first TNFα cycle. No complete hematologic remission or cytogenetic improvement was seen. Side-effects were similar to those described for both substances alone. Maximum tolerable TNF doses usually varied between 80 µg/m2 and 160 µg/m2. To examine possible pathways of TNF activity in these patients, interferon receptor status and (2′–5′)-oligoadenylate synthetase levels were examined in peripheral blood mononuclear cells. Both parameters remained unchanged during TNFα treatment. These preliminary data point to significant clinical efficacy of additionally applyed TNFα in IFNα-resistant CML patients.

Analysis of the novel cyclin-dependent kinase 4 and 6 inhibitor gene p18 in lymphoma and leukemia cell lines

The genes for the CDK4/6-inhibitors p16INK4A/MTS1 and p15INK4B/MTS2 are frequently deleted in hematological malignancies. A new member of this family of CDK4/6 inhibitors is p18. In order to assess p18 growth-suppressor gene alterations in hematological neoplasms, we investigated 31 lymphoma and leukemia cell lines by PCR for both exons of this gene. No homozygous deletions were observed. Investigation of a new intragenic restriction fragment length polymorphism revealed no differences in allele distribution between the tumor cell lines and healthy volunteers. Our results suggest that homozygous deletion of the p18 gene does not play a major role in leukemogenesis or lymphomagenesis.

DNA-flow cytometry studies in blood and marrow cells from chronic myelogenous leukemia patients treated with interferon alpha-2b

Cell cycle distribution in bone marrow and peripheral blood mononucleated cells was studied in patients with chronic myelogenous leukemia (CML) before and during treatment with interferon (IFN) alpha-2b. DNA-flow cytometry with ethidium bromide fluorescence was used. Highly significant differences between mononucleated cells from CML patients and normal controls were seen in peripheral blood but not in bone marrow specimens. Patients achieving hematologic remission during IFN treatment showed a cell cycle distribution in bone marrow cells and peripheral blood cells similar to normal controls.

Ifosfamide. Methotrexate and 5-fluorouracil for pretreated advanced breast cancer

A total of 51 fully evaluable patients with advanced and intensively pretreated breast cancer were treated with a combination chemotherapy of ifosfamide plus mesna, methotrexate and 5-fluorouracil. All patients had received at least one series of combined chemotherapy, 30 patients had received more than one combination and 41 patients had had anthracyclines before. Metastatic lesions in more than one site were found in 42 patients, and 24 patients had metastatic liver lesions. Partial remission was achieved in 10 patients (20%) and no change in 16 patients (31%). Survival was almost identical in both groups of responding patients and significantly shorter in treatment failures. Response was favorable in patients without pretreatment with anthracyclines. Two patients who received this protocol directly after progression with cyclophosphamide, methotrexate and 5-fluorouracil (CMF protocol) responded with a partial remission. Median time to progression was 7 months for partial responders and 4.5 months for patients achieving a no-change status. Median survival was 8 months for all patients. Toxicity was tolerable. Leukocytopenia and thrombocytopenia were treatment-limiting parameters. Overall, this protocol is well tolerable and effective in breast cancer patients with advanced disease and in intensively pretreated patients.

RAPAMYCIN ANTAGONIZES INTERLEUKIN-6 MEDIATED GROWTH ARREST AND DIFFERENTIATION OF MYELOBLASTIC M1 CELLS

The immunosuppressant rapamycin is known to be a potent inhibitor of cytokine driven proliferation of T-lymphocytes and other cell types. Here we have examined the effect of rapamycin on the myeloblastic cell line M1 which responds to interleukin-6 (IL-6) with growth arrest and monocytoid differentiation. Single-agent rapamycin led to a retardation of M cell growth. In spite of this intrinsic antiproliferative effect, rapamycin was found to abrogate IL-6 induced growth arrest. Concomitant exposure to rapamycin and IL-6 also reduced the extent of monocytoid differentiation as compared to treatment with IL-6 alone. Excess levels of the FK-506 analogue ascomycin reversed the antagonistic effect of rapamycin on IL-6 mediated growth suppression, suggesting that this biological action of rapamycin is mediated by a rapamycin/immunophilin complex. The findings demonstrate that rapamycin can also act as an antagonist of cytokine induced growth arrest and differentiation.