Chronic myelogenous leukemia: recent developments in prognostic evaluation and chemotherapy. The German CML Study Group.

Recent developments in CML research are illustrated by the results of one large randomized multicenter study carried out by the German CML Study Group. From July 1983 to January 1991, a total of 703 CML patients were recruited; 624 patients were randomized to compare hydroxyurea and interferon alpha (IFN) with busulfan. The median survival of Ph+ patients by now is 3.95 years, that of Ph- patients 1.1 years. Some difference in survival is recognizable between the treatment arms, but this is not yet significant. Fewer adverse effects are being observed in the hydroxyurea group. Ph-negative patients tend to have lower white blood cell and platelet counts. Patients (164) were randomized to receive IFN. In 50 patients (30%) IFN had to be terminated because of adverse effects, therapy resistance, or other reasons. Reduction of the Ph-chromosome was observed in 20% of evaluable patients. In 3 patients complete cytogenetic remissions were observed. Clinically relevant neutralizing antibodies were detected in 9 cases. Prospectively evaluated age, organomegaly related symptoms, Karnofsky index, extramedullary manifestations, erythroblasts, and percent of circulating blasts proved to be of prognostic significance. A prognostic score (score 1) was determined and compared to Sokal's score. It is expected that the study results will allow statements as to the advantages or disadvantages of the use of busulfan, hydroxyurea and IFN in the treatment of CML as well as to the reliability of prognostic markers.

The German CML study, comparison of busulfan vs. hydroxyurea vs. interferon alpha and establishment of prognostic score 1.

From July 1983 to January 1991 a total of 622 patients were randomized (585 eligible) to compare the effects of hydroxyurea, interferon alpha (IFN), and busulfan on the duration of chronic phase, and survival. Further goals included the determination of prognostic parameters. 598 CML patients were documented and 575 evaluable. The Ph-status was known for 547 patients. 89.4% of the patients were Ph-positive (+). 11% had additional chromosome aberrations. The median survival of Ph+ patients by now is 4.2 years, that of Ph-patients 1.4 years. Ph-negative patients are older, tend to have lower cell counts and, as a group are more ill at diagnosis. A survival difference of about one year is expected between busulfan and hydroxyurea treated patients. Prospectively evaluated age, organomegaly related symptoms, Karnofsky index, extramedullary manifestations, number of erythroblasts and percent of circulating blasts proved to be of prognostic significance. A prognostic score (score 1) was determined which was superior to Sokal's score in the study population. 164 patients were randomized to receive IFN. In 54 patients (33%) IFN had to be terminated because of adverse effects, therapy resistance or other reasons. Clinically relevant neutralizing antibodies were detected in 9 cases. Most frequent adverse events were flu-like symptoms in 74%, gastrointestinal symptoms in 52%, and neurologic-psychiatric symptoms in 30% of patients. Reduction of the Ph-chromosome was observed in 13% of evaluable patients (10 of 75). In 4 patients complete cytogenetic remissions were observed, in three of these ongoing. Cytogenetic responders have a survival advantage. Interferon treated Philadelphia-negative CML patients have no survival disadvantage. The study is expected to allow statements as to the advantages or disadvantages of the use of busulfan, hydroxyurea and IFN in the treatment of CML as well as to the reliability of prognostic markers.

[Erythropoietin assay. Comparison of a biological test system with an haemagglutination inhibition test (author's transl)]. / Nachweis von Erythropoietin. Vergleich eines biologischen Verfahrens mit einem Haemagglutinations-Hemmtest.

The results from a biological test for erythropoietin (using the rate of iron absorption in polycythemic mice) and a commercially-available immunological test (haemagglutination-inhibition test) were compared. Of 19 batches of the immunological test which were investigated, 7 batches were completely inactive and a further 3 batches reacted only with the test serum supplied with the test. There was a poor correlation between the results from the biological and the immunological measurements, both on patients with high and those with low serum erythropoietin levels. The difficulty of the immunological erythropoietin test is that pure erythropoietin is not sufficiently available. The immunological test investigated here does not use pure erythropoietin. Aside from this, pathophysiological considerations would lead one to expect basic differences between the results from immunological and biological tests.

[On the existence of a myeloproliferative factor in patients with a myeloproliferative syndrome (author's transl)]. / Zur Frage der Existenz eines myeloproliferativen Faktors bei Patienten mit myeloproliferativem Syndrom.

Serum of patients suffering from a chronic myeloproliferative disorder (polycythaemia, era, osteomyelofibrosis, chronic myeloid leukaemia) and serum of lethally irradiated rats injected before application of a single doses of erythropoietin did not enhance the effect of erythropoietin -- measured with the iron incorporation rate of polycythemic mice. The rationale for these experiments is to try to find a "myeloproliferative factor", which augments the number of stem cells as described in sera of patients with polycythaemia vera, osteomyelofibrosis, and lethally irradiated mice.

[Spontaneous lymphocyte transformation in chronic renal insufficiency (author's transl)]. / Spontane Lymphocytentransformation bei chronischer Niereninsuffizienz

3H-thymidine uptake of lymphocytes cultured from patients with regular dialysis treatment was determined at the time of maximal transformation, which was ascertained in previous investigations to be 5 days after the beginning of incubation. Spontaneous DNS synthesis was found to be significantly depressed: 1137 plus or minus 1122 c.p.m./culture; controls: 9783 plus or minus 7499 c.p.m./culture, whereas LDH activity in the supernatant culture medium measured as a parameter of continuous cell destruction showed no alteration. After stimulation with 400 mug PHA DNS synthesis in 7 patients was also decreased, but elevated in 5 patients with previous infections. After incubation with added serum from 11 chronic uremic patients, spontaneous lymphocyte transformation in cultures sampled from 2 healthy subjects was also found to be depressed with positive correlation to the hemoglobin and hematocrit levels in the corresponding blood. The significantly increased LDH activity values in the medium as the consequence of impaired cll viability showed also positive correlation with 3-H-thymidine uptake levels; therefore depressed DNS synthesis cannot be due alone to accelerated cell destruction. In analogy to the erythrocytes, two stimultaneous effects of uremic toxins in serum are assumed to be exerted on the lymphocytes: metabolic inhibition of DNS synthesis and impaired cell viability.

Large-scale isolation of middle and higher molecular weight uremic toxins.

Reverse osmosis (40 bar) using membranes with a nominal cut-off of 500 Dalton is a useful method for the desalting and concentration of hemofiltrates (20-30 L) or even dialysates (120-240 L) from patients with end-stage renal failure. The removal of electrolytes and lower molecular weight solutes from the middle and higher molecular weight fractions can be carried out in one step. The freeze-dried residues show characteristic differences in their molecular weight distributions which are dependent upon their origin (Cuprophane dialysates or RP-6-hemofiltrates). Subfractionation is performed using Sephadex G-15 macrocolumns (2 m x 5 cm), the LKB-Ultrogel AcA 54 and Sephacryl S-200 as well as ion exchange chromatography. The fractions are characterized by their ability to suppress the incorporation of 3H-thymidine into rat bone-marrow as well as HeLa cell cultures. The greatest inhibitory activity originates from those fractions which are not dialysable using Visking tubes in vitro. The results indicate that the middle and higher molecular weight spectra do in fact overlap, suggesting that also higher molecular weight substances are involved in uremic intoxication.

[Long-term cultures in polycythaemia vera as proof for dependence on erythropoietin (author's transl)]. / Langzeitkulturen bei Polycythaemia vera zum Nachweis von Erythropoetinabhängigkeit.

In eight patients suffering from polycythaemia vera, the proliferation of erythroid colonies in methylcellulose with and without exogenous erythropoietin was studied. After three and five weeks, respectively, no colonies grew. Due to addition of erythropoietin more erythroid colonies proliferated which could be cultivated over nine weeks. The experiments suggest that the erythropoiesis in polycythaemia vera depends on erythropoietin.

Preparative isolation of middle molecular weight fractions from the hemofiltrate of patients with chronic uremia.

Fractions in the molecular weight range of the so-called middle molecules (500--3000 daltons) were isolated on a preparative scale from hemofiltrate of patients. Hemofiltration was performed with an RP-6 dialyzer using a predilution technique. The hemofiltrate was concentrated and desalinated by reverse osmosis with membranes with a normal cut-off of 500 daltons. The retentate was freeze-dried, redissolved in volatile buffer and fractionated on Sephadex G-15 macrocolumns. The middle molecule molecular weight range in the elution profiles (detected at 206 and 280 nm simultaneously) was marked with peptides. Isolated fractions in the middle molecule molecular weight range showed strong inhibitory activity upon 3H-thymidine incorporation into rat bone-marrow cells in vitro. The described combination of hemofiltration, reverse osmosis and gel chromatography is suggested as a useful approach to the isolation of these ninhydrin-positive substances which, according to their proved heterogeneity, are to be subfractionated by further procedures.

[Effect of higher molecular urine metabolites on lymphocyte transformation and autohemolysis (author's transl)]. / Lymphozytentransformation und Autohämolyse unter Einwirkung höhermolekularer Urinmetabolite als potentielle Urämietoxine

Possible toxic effects of lyophilised not dialysable residue from urine of healthy persons obtained by 4 days dialysing against water were investigated. Therefore, the substance dissolved at a concentration of 2.22 mg/ml in Medium TC 199 respectively in physiological saline solution was added with increasing quantities (0.1 to 1.0 ml) to lymphocyte cultures (3 x 10(6) cells in 4 ml medium) and to erythrocytes (3 ml of blood after separation of granulocytes) from healthy persons. Spontaneous 3H-thymidine uptake of the lymphocytes ascertained 5 days after beginning of incubation was markedly depressed with dependence upon the doses up to 30.4% of the control value, whereas only 65.2% was reached after stimulation with 400 mug PHA. In both cases LDH activity in the supernatant culture medium measured as a parameter of cell destruction showed a dose-dependent increase with inverse trend as compared to 3H-thymidine uptake levels. This negative correlation may be due to impaired cell viability as the main cause for depressed lymphocyte transformation. The autohemolysis of the erythrocytes was diminished with dependence upon the doses until to 59.3% of the control value. Therefore, only cytotoxic effects of uremic serum on the lymphocytes may be due to higher molecular urine metabolites retained in uremia whereas increased autohemolysis will be induced by toxins of lower molecular weight.

Recombinant human erythropoietin in transfusion-dependent anemic patients with multiple myeloma and non-Hodgkin's lymphoma--a randomized multicenter study. The European Study Group of Erythropoietin (Epoetin Beta) Treatment in Multiple Myeloma and Non-Hodgkin's Lymphoma.

One hundred twenty-one anemic, transfusion-dependent patients with multiple myeloma (MM) or low-grade non-Hodgkin's lymphoma (NHL) were randomly allocated to receive (a) recombinant human erythropoietin (rhEPO) 10,000 U/d subcutaneously 7 days a week (fixed dose group) (n = 38), or (b) rhEPO 2,000 U/d subcutaneously for 8 weeks followed by step-wise escalation of the rhEPO dose (titration group) (n = 44), or (c) no rhEPO therapy (control group) (n = 39). The total treatment period was 24 weeks. There were no differences between the three groups with regard to baseline clinical, demographic, or health status measures. The cumulative response frequency, defined as elimination of the transfusion need in combination with an increase in the hemoglobin concentration by >20 g/L, was 60% in both rhEPO treatment groups and 24% in the control group (P = .01 and .02, respectively, log rank test). For patients in the titration group the response rate on the first dose level (2,000 U/d) was only 14%. Cox's univariate regression analysis revealed that an inadequately low endogenous erythropoietin concentration in relation to the degree of anemia and a baseline platelet concentration > or = 100 x 10(9)/L were significant predictors for response to rhEPO therapy (P < .01). Multivariate regression analysis showed that relative erythropoietin concentration was the most important factor and the platelet count had no additional influence on response. Treatment with rhEPO was well tolerated. We conclude that treatment with rhEPO may be indicated in anemic MM and NHL patients with a relative erythropoietin deficiency. An initial dose of 5,000 U/d subcutaneously may be recommended.

Phase II study of pirarubicin in metastatic breast cancer.

Pirarubicin is a more lipophilic derivative of doxorubicin, with a higher uptake rate of cells, lower cardiotoxicity and better antitumor efficacy in preclinical models. Thirty-four patients with metastatic breast cancer were treated in a multicenter phase II study with pirarubicin (THP) using a dosage of 75 mg/m2/every 3 weeks. The patients had a median age of 56 years (range 41-73) and a performance status of WHO grade 0-2. Patients pretreated with anthracyclines, or who were older than 75 years and without sufficient bone marrow reserve were excluded. The 32 evaluable patients received a median number of 4 cycles (range 2-8). The myelosuppression was dose-limiting and led to infections (grades 1 and 2) in 5 patients. Twenty-eight patients developed leukocytopenia grade 3 and 4 toxicity and 7 patients experienced thrombocytopenia grade 1 and 2. The drug was subjectively well tolerated and nausea, vomiting and alopecia were mild. One complete remission with a duration of 15.4 months (67 weeks) and 7 partial remissions with a median duration of 9.3 months (40 weeks) were achieved, which resulted in an overall response rate of 25%. Twenty-one patients were stable for 17 weeks (median) under the treatment with pirarubicin.