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.

Azacitidine in combination with intensive induction chemotherapy in older patients with acute myeloid leukemia: The AML-AZA trial of the Study Alliance Leukemia.

DNA methylation changes are a constant feature of acute myeloid leukemia. Hypomethylating drugs such as azacitidine are active in acute myeloid leukemia (AML) as monotherapy. Azacitidine monotherapy is not curative. The AML-AZA trial tested the hypothesis that DNA methyltransferase inhibitors such as azacitidine can improve chemotherapy outcome in AML. This randomized, controlled trial compared the efficacy of azacitidine applied before each cycle of intensive chemotherapy with chemotherapy alone in older patients with untreated AML. Event-free survival (EFS) was the primary end point. In total, 214 patients with a median age of 70 years were randomized to azacitidine/chemotherapy (arm-A) or chemotherapy (arm-B). More arm-A patients (39/105; 37%) than arm-B (25/109; 23%) showed adverse cytogenetics (P=0.057). Adverse events were more frequent in arm-A (15.44) versus 13.52 in arm-B, (P=0.26), but early death rates did not differ significantly (30-day mortality: 6% versus 5%, P=0.76). Median EFS was 6 months in both arms (P=0.96). Median overall survival was 15 months for patients in arm-A compared with 21 months in arm-B (P=0.35). Azacitidine added to standard chemotherapy increases toxicity in older patients with AML, but provides no additional benefit for unselected patients.

Treatment strategies in patients with AML or high-risk myelodysplastic syndrome relapsed after Allo-SCT.

Non-relapse mortality after Allo-SCT has significantly decreased over the last years. Nevertheless, relapse remains a major cause for post SCT mortality in patients with AML and high-risk myelodysplastic syndrome (MDS). In this retrospective single-center analysis, we have analyzed the treatment outcomes of 108 patients with AML or MDS, who relapsed after Allo-SCT. Seventy of these patients (65%) were treated with salvage therapies containing chemotherapy alone, allogeneic cell-based treatment or the combination of both. Thirty-eight patients (35%) received palliative treatment. Median OS after diagnosis of relapse was 130 days. Compared with patients who received chemotherapy alone, response to salvage therapy was significantly improved in patients treated with a combination of chemo- and allogeneic cell-based therapy (CR rate 57% vs 13%, P=0.002). Among risk factors concerning pretreatment characteristics, disease status before first Allo-SCT, and details of transplantation, only the time interval from Allo-SCT to relapse was an independent predictor of response to salvage therapy and OS. These data confirmed that time to relapse after transplantation is an important prognostic factor. Up to now, only patients eligible for treatment regimens containing allogeneic cell-based interventions achieved relevant response rates.

CD34(+) lineage specific donor cell chimerism for the diagnosis and treatment of impending relapse of AML or myelodysplastic syndrome after allo-SCT.

After allo-SCT, analysis of CD34(+) lineage-specific donor cell chimerism (DCC) is a sensitive method for monitoring minimal residual disease in patients with AML or myelodysplastic syndrome (MDS) with CD34 expression. To substantiate evidence of whether immune interventions in patients with impending relapse, defined by incomplete lineage-specific DCC, may prevent hematological relapse, we performed a retrospective nested case control study. Unsorted and lineage-specific DCC were measured in 134 patients. Forty-three patients had an incomplete CD34(+)-DCC with no other evidence of relapse. After immediate tapering of immunosuppressive treatment (30 patients) and/or infusion of donor lymphocytes (10 patients), 21 patients remained in remission (conversion to complete lineage-specific DCC) and 22 relapsed. Relapse-free survival at 3 years of the 91 patients with stable DCC and of the 43 patients with incomplete DCC was 74% (95% confidence interval (CI), 64-83%) and 40% (95% CI, 24-58%), respectively. OS rates were 79% (95% CI, 70-88%) and 52% (95% CI, 35-69%), respectively. These results, with 49% of patients with impending relapse successfully treated with immune intervention, highly suggest that analysis of CD34(+)-DCC is an important tool for monitoring and the management of AML and MDS patients after allo-SCT.

High-dose chemotherapy with autologous PBSC transplantation for poor prognosis germ cell tumors: a retrospective monocenter analysis of 44 cases.

Germ cell cancer (GCC) is curable in metastatic stages. The International Germ Cell Cancer Collaborative Group (IGCCCG) reports a poor prognosis subgroup with a 5-year survival of 48%. High-dose chemotherapy with PBSC transplantation (HD-PBSCT) in these patients showed promising results in phase II, but failed to show significant advantage in randomized trials. We report our monocenter series of all poor and selected intermediate prognosis germ cell tumor patients treated with multiple-course HD-PBSCT and secondary surgery of remaining tissue. We performed a retrospective analysis of our complete series of 44 patients (40 poor prognosis and 4 intermediate prognosis) treated by HD-PBSCT as part of first-line therapy from 1999 to 2010. The CR rate after up to four cycles of HD-PBSCT and radical resection of residual manifestations was 73%. The 3-year survival rate was 79.5% (median follow-up of 51.5 months; range: 7-143 months). Disease-related death rate was 16%. HD-PBSCT-related death did not occur. One patient died postsurgery. Multiple courses of HD-PBSCT with radical secondary surgery is safe and effective in poor prognosis metastatic GCC. Despite disappointing phase III studies it is of high interest to further study this field.

Induction therapy of AML with ara-C plus daunorubicin versus ara-C plus gemtuzumab ozogamicin: a randomized phase II trial in elderly patients.

BACKGROUND: Chemotherapy for elderly patients with acute myeloid leukemia (AML) results in a median overall survival (OS) of ≤ 1 year. Elderly patients often present with cardiac comorbidity. Gemtuzumab ozogamicin (GO) is active in elderly (≥ 60 years) patients with relapsed AML with low cardiac toxicity. PATIENTS AND METHODS: This randomized phase II study compared a standard combination of ara-C and daunorubicin (DNR; 7+3) versus ara-C plus gemtuzumab ozogamicin (7+GO) as the first course of induction therapy. Primary objectives were comparison of blast clearance on day 16, event-free survival (EFS), and remission duration. OS, complete remission (CR), and tolerability were secondary objectives. RESULTS: One hundred and nineteen patients with de novo AML, treatment-related AML, AML with a history of myelodysplastic syndrome (MDS), or high-risk MDS entered the study. Median age of 115 patients (intent-to-treat population) was 69 years. Protocol outlined a second course 7+3 for patients without blast clearance and two courses of high-dose ara-C consolidation upon CR. Both treatments were equally effective in blast clearance, CR, EFS, remission duration, or OS (median: 7+3, 9 months; 7+GO, 10 months). Induction death rate was higher in the GO group due to veno-occlusive disease. CONCLUSION: The study did not show significant superiority of 7+GO over standard 7+3.

Amsacrine containing induction therapy in elderly AML patients: comparison to standard induction regimens in a matched-pair analysis.

Many elderly patients with newly diagnosed acute myeloid leukemia (AML) present with cardiac comorbidity precluding the use of anthracycline containing chemotherapy regimens. Amsacrine, a topoisomerase II inhibitor, has been proposed as possible alternative to anthracyclines. Here, we report about the combination of amsacrine (210 mg/m(2)), in replacement for daunorubicin (DNR), with standard dose cytarabine and thioguanine (TAA) to elderly patients (>or=60 years of age) with impaired cardiac function. The outcome of 16 patients with a median age of 66 years treated between 1997 and 2003 was compared with standard treatment regimens of the AMLCG study group in a matched-pair analysis. There were no statistically significant differences in response rate, relapse free survival or overall survival between TAA treated patients or standard therapy. In conclusion, replacing anthracyclines with amsacrine for induction therapy of AML patients with significant cardiac comorbidities represents a treatment option without compromising the potential curability of the disease.

The emergence of a C/EBPalpha mutation in the clonal evolution of MDS towards secondary AML.

Recently, mutations in the transcription factor CCAAT/ enhancer binding protein alpha (C/EBPalpha) have been described in acute myeloid leukemia (AML). We performed a mutational analysis of the C/EBPalpha gene in the myelodysplastic syndromes and AML with antecedent MDS. No mutations were found in patients with refractory anemia (0/27), refractory anemia with ringed sideroblasts (0/7), refractory anemia with excess of blasts (RAEB 0/16) or chronic myelomonocytic leukemia (CMML 0/5). One out of 13 patients with RAEB-T/AML secondary to MDS showed a mutation in the C/EBPalpha gene. In this patient a 4 bp insertion disrupted codon 69 in one allele. This novel +1 frame shift is predicted to result in a truncated protein of 107 amino acids. However, the dominant protein translated was the C/EBPalpha isoform p30, which was previously shown to inhibit the DNA-binding and transactivation properties of C/EBPalpha p42. Interestingly this mutation could not be detected at diagnosis in the initial RAEB and RAEB-T stage. The mutation appeared at relapse after chemotherapy for RAEB-T. We conclude that the C/EBPalpha mutation was not essential for the initial blast accumulation. The emergence of a bast clone carrying a C/EBPalpha mutation at relapse indicates that this mutation may confer a growth advantage in a myeloid cell with an established differentiation block.

Real-time RT-PCR for the detection and quantification of AML1/MTG8 fusion transcripts in t(8;21)-positive AML patients.

AML1/MTG8 was quantified relative to the expression of the GAPDH housekeeping gene by real-time RT-PCR in 22 patients with t(8;21)-positive acute myeloblastic leukaemia (AML) at initial diagnosis and in seven of these patients also during/after chemotherapy and allogeneic bone marrow transplantation. Real-time PCR was able to specifically detect and quantify AML1/MTG8 over a 5 log range. The detection limit for t(8;21)-positive cells was a dilution of 1:105. The AML1/MTG8 expression varied considerably among the 22 AML patients at intial diagnosis with a ratio AML1/MTG8:GAPDH of 0.5135+/-0.536 (range 0.1-2.14, median 0.318). In six patients with t(8;21)-positive AML a marked decline of AML1/MTG8 could be induced by chemotherapy. These patients are in ongoing complete haematological remission (CR) with a constant low-level AML1/MTG8 expression. In another patient a rapid rise of AML1/MTG8 transcripts could be detected in CR after allogeneic bone marrow transplantation and the patient relapsed 10 weeks later. In conclusion, real-time RT-PCR is a suitable approach for the quantification of AML1/MTG8 transcripts in the monitoring of AML patients with t(8;21) during/after chemotherapy and can provide data of prognostic relevance.

Migraine associated bilateral intracerebral haemorrhages.

The authors report a case of bilateral basal ganglionic haemorrhages which occurred during an attack of classical migraine. The patient had a history of migraine associated with aura of neurological deficit for 10 years and a history of arterial hypertension for 20 years, which was treated with propranolol. Intracerebral haemorrhage during an attack of migraine is very rare and up to now the existence of true migraine-induced intracerebral haemorrhage has been controversial. Our case of bilateral occurrence of the haemorrhages supports the theory of the existence of migraine-induced damage of the wall of intraparenchymal vessels during vasoconstriction and focal ischaemia at the beginning of a migraine attack. Subsequent vessel rupture may occur during the following period of increased cerebral blood flow especially with coexisting arterial hypertension. The terminology of the syndrome of migraine associated with intracerebral haemorrhage is reviewed.

Modulation of human glucokinase intrinsic activity by SH reagents mirrors post-translational regulation of enzyme activity.

The low-affinity glucose phosphorylating enzyme glucokinase plays a key role in the process of glucose recognition in pancreatic B-cells. To evaluate mechanisms of intrinsic regulation of enzyme activity human pancreatic B-cell and liver glucokinase and for comparison rat liver glucokinase were expressed in E. coli bacteria. A one-step purification procedure through metal chelate affinity chromatography revealed 58 kDa proteins with high specific activities in the range of 50 U/mg protein and K(m) values around 8 mM for the substrate D-glucose with a preference for the alpha-anomer. There were no tissue specific differences, no species differences in the electrophoretic mobility, and no differences of the kinetic properties of these well conserved enzymes. The deletion of the 15 tissue-specific NH2-terminal amino acids of the human glucokinase resulted in a catalytically active enzyme whose kinetic properties were not significantly different from those of the wild-type enzymes. The human and rat glucokinase isoforms were non-competitively inhibited by the sulfhydryl group reagents alloxan and ninhydrin with Ki values in the range of 1 microM. The inhibition of glucokinase enzyme activity was reversed by dithiothreitol with an EC50 value of 9 microM for alloxan and of 50 microM for ninhydrin. D-Glucose provided protection against alloxan-induced inhibition of human and rat glucokinase isoenzymes with half-maximal effective concentrations between 11 and 16 mM. The enzyme inhibition by alloxan was accompanied by a change in the electrophoretic mobility with a second lower molecular 49 kDa glucokinase band which can be interpreted as a compact glucokinase molecule locked by disulfide bonds. Quantification of free sulfhydryl groups revealed an average number of 3.6 free sulfhydryl groups per enzyme molecule for the native human glucokinase isoforms. Alloxan decreased the average number of free sulfhydryl groups to 1.9 per enzyme molecule indicating that more than one SH side group is oxidized by this compound. The extraordinary sensitivity of the SH side groups of the glucokinase may be a possible mechanism of enzyme regulation by interconversion of stable (active) and unstable (inactive) conformations of the enzyme. In pancreatic B-cells the glucose-dependent increase of reduced pyridine nucleotides may stabilize the enzyme in the 58 kDa form and provide optimal conditions for glucose recognition and glucose-induced insulin secretion.

[Changes in the cornea following complicated pseudophakia and subsequent keratoplasty (clinical and scanning electron microscope findings)]. / Veränderungen der Hornhaut nach komplizierter Pseudophakie und anschliessender Keratoplastik (klinische und REM-Befunde).

An anterior chamber lens (Binkhorst four-loop lens) became dislocated seven weeks after implantation. It was relocated in the right position by means of a small incision. In the course of the following 8 weeks the cornea opacified from the top downward. A penetrating keratoplasty was performed. Intraoperatively it was found that a strand of vitreous had pushed forward between the iris and the lens and was adhering to the back of the cornea. The piece of cornea removed was prepared for the scanning electron microscope. The microscopic findings are described in detail. They correspond exactly to the clinical diagnosis of complete tearing of the endothelium, considerable bullous changes in the parenchyma and degenerative changes in the epithelium. The scanning electron microscope reveals how severe pathologic changes in the cornea can become if a strand of the vitreous becomes adherent to the endothelium.

[Etiology and pathogenesis of recurrent erosions of the cornea. A scanning and transmission electron microscopy study]. / Atiologie und Pathogenese der rezidivierenden Erosion der Hornhaut. Eine raster- und transmissionselektronenmikroskopische Studie.

Scratched off corneae of 2 patients were examined under the scanning electron and the transmission electron microscope. Whereas in the scanning electron microscope the expected picture of the destruction of the epithelium in the upper zones of the cornea could be presented in the 1st case, the histological picture of the 2nd case shows in the transmission electron microscope the reaction of the influence of the noxa to the texture of the epithelial cells, to the protoplasm, and to the nucleus of the concerned epithelial cell.

[Electron-microscopic examinations in cases of keratitis dendritica (author's transl)]. / Elektronenmikroskopische Untersuchungen bei Keratitis dendritica.

Scanning electron-microscopic and transmission electron-microscopic examination of cornea affected with herpes simplex (DNS) and which developed the clinical picture of keratitis dendritica, showed typical test results, which are described in detail. The ultramicroscopic findings show that the metabolism of the epithelial cells is of considerable degree. It was proved that there were virus particles in the protoplasm of the epithelial cells. A comparison with a case of zoster keratitis showed the same morphological changes.

[The "typical senile cataract" examined under the scanning electron microscope (author's transl)]. / Der "typische graue Altersstar" im Rasterelektronen mikroskop (REM).

Lenses suffering from "typical senile cataract" prepared by "critical point drying"and by sprinkling with gold, were examined under the scanning electron microscope. The images thus obtained gave a three-dimensional representation of structural changes within the lens determined in histological investigations performed with the light microscope.

Emergence of insulin receptors on human lymphocytes during in vitro transformation.

Essentially no specific binding sites for insulin are detected in small lymphocytes freshly isolated from human blood. Insulin-binding sites appear on the lymphocyte surface during transformation in vitro with concanavalin A, and the number of these receptors increases sharply to reach a maximum between 24 and 46 hr after exposure to the mitogen. The appearance of de novo binding sites for insulin coincides with the increase in [(3)H]thymidine uptake into nuclear DNA and clearly precedes the appearance of enlarged, morphologically transformed cells. No changes in insulin-binding are detected in unstimulated control cultures. A maximum of about 350 molecules of insulin can bind per transformed lymphocyte, while less than six insulin molecules bind to an untransformed cell. Circulating human leukemic lymphoblasts bind about as much insulin as the lymphocytes transformed in vitro. Giant, polynucleated, transformed lymphocytes cultured in the presence of cytochalasin B bind about 10 times more insulin than transformed lymphocytes, which is in harmony with a 10-fold increase in cell-surface area in these cells. Specific binding of insulin is a saturable process in transformed lymphocytes but not in the untransformed cells. In transformed cells, [(125)I]-insulin is displaced by as little as 2 ng/ml of native insulin, while in untransformed cells no significant displacement is observed with native insulin. Digestion of transformed cells with phospholipase C (EC 3.1.4.3.) enhances the specific binding of [(125)I]insulin 3-fold, but no effect occurs with untransformed cells. These observations indicate a possible functional role of insulin and of adenylate cyclase in cell growth and division.