Sequence of administration and methylation of SOCS3 may govern response to gemtuzumab ozogamicin in combination with conventional chemotherapy in patients with refractory or relapsed acute myelogenous leukemia (AML).

In older patients suffering from acute myelogenous leukemia (AML), aggressive chemotherapy is accompanied with high treatment-related morbidity and mortality. Gemtuzumab ozogamicin (GO), a humanized monoclonal anti-CD33 antibody, represents a well tolerated treatment option, but optimal treatment schedules are still unknown. Additionally, Suppressor of cytokine signaling 3 (SOCS3) inhibits the CD33-induced block on cytokine-induced proliferation. Consequently, a variable response of AML cells to anti-CD33-targeted therapy may be caused by modulation of SOCS3 expression. Twenty-four patients with refractory or relapsed CD33-positive AML received GO as a single agent before or after conventional chemotherapy. The methylation status of the SOCS3 CpG island was assessed by methylation-specific polymerase chain reaction. Response (RR) and overall survival (OS) were significantly higher in 16 patients receiving chemotherapy before GO (RR 81%, OS 14.8 months) compared to three patients who received GO single agent therapy (RR 33%, OS 7.2 months) or 16 with GO before chemotherapy (RR 0% OS 2.2 months, P = 0.01 for RR and P < 0.001 for OS). Methylation of the SOCS3 CpG island was found in 8/24 patients. There was a trend towards a higher RR and longer OS in patients with SOCS3 hypermethylation (RR 86%, OS 25.1 months) compared to unmethylated SOCS3 (RR 56%, OS 10.3 months, P = 0.09). Administration of GO a few days after chemotherapy seems to provide better response and survival compared to administration of GO directly before chemotherapy. The potential role of SOCS3 hypermethylation as a biomarker should be further investigated in patients undergoing GO containing therapies.

Antineoplastic chemotherapy in cancer patients with methicillin-resistant Staphylococcus aureus (MRSA).

BACKGROUND: Methicillin-resistant Staphylococcus aureus (MRSA) is a major nosocomial pathogen causing serious morbidity and mortality in immunosuppressed patients. Antineoplastic chemotherapy causes immunosuppression, and thus there is concern whether such patients should proceed to therapy without delay or dose reduction. There are presently no guidelines with appropriate provisions for antineoplastic chemotherapy in cancer patients with MRSA colonization or infection. PATIENTS AND METHODS: We retrospectively analyzed the clinical outcome of all 27 patients with known MRSA infection or colonization undergoing antineoplastic chemotherapy for solid or hematological malignancies in our institution. RESULTS: In our patients, MRSA was detected at multiple sites. 11 patients were found to be colonized with MRSA, whereas 16 patients had colonization and/or infection. MRSA sepsis occurred in 12 cases. Interestingly, at the time of MRSA sepsis, neutrophil counts were less than 500/µl in 42% of our patients. However, fatal complications due to MRSA occurred in only 2 patients. Among patients with MRSA sepsis, the mortality rate was 14%. CONCLUSIONS: Our results with a limited number of patients support the contention that antineoplastic chemotherapy may well be administered to patients with MRSA and should not necessarily lead to dose reduction or treatment delay, especially in cases with curative intent.

Successful treatment of hepatic encephalopathy in a patient with acute lymphoblastic leukemia.

Infiltration of the liver with consecutive severe dysfunction is rarely seen in adult acute lymphoblastic leukemia (ALL). We describe a case of a 32-year-old woman with severe icterus, thrombocytopenia, neutropenia and hepatosplenomegaly. ALL was diagnosed. One day after admission, she developed hepatic encephalopathy with ammonia concentrations in plasma >100 micromol/l. Hepatic infiltration was presumed and chemotherapy was initiated immediately which led to resolution of hepatic encephalopathy and complete hematological remission. Clinicians should be aware of unusual presentations of ALL and difficulties for the application of chemotherapy in patients with liver failure.

Cisplatin as single-agent chemotherapy in patients with liver dysfunction due to metastases.

BACKGROUND: Chemotherapeutic options are often limited for patients with hepatic dysfunction induced by advanced metastases. The toxicity and efficacy of cytotoxic drugs are often unpredictable due to altered drug activation or inactivation and excretion. Therefore, numerous chemotherapeutic agents should not be administered to patients with liver failure. PATIENTS AND METHODS: We retrospectively analysed 14 patients with solid tumours presenting with liver dysfunction induced by metastasis who were treated with cisplatin single-agent chemotherapy. RESULTS: Tolerance of therapy was acceptable with 1 grade I and 1 grade II renal toxicity. 6 patients experienced grade III haematological toxicity. Partial remission of the disease was observed in 6 cases, and 6 patients could receive combination chemotherapy after improvement of liver function. The median overall survival of the patient cohort was 4.8 months. CONCLUSION: Cisplatin monotherapy may thus be considered as a treatment option for patients with liver dysfunction induced by different solid tumours.

Molecular evidence for a clonal relationship between multiple lesions in patients with unknown primary adenocarcinoma.

Unknown primary adenocarcinoma (UPA) comprises a group of heterogeneous cancers of great clinical and biological interest. UPA presents as metastatic disease without a detectable primary site after medical workup. Here we investigated whether or not a clonal relationship exists between multiple tumors within individual UPA patients. A molecular resemblance would argue for an early clonal outgrowth of tumor cells from the primary lesion, a mutual feature observed within this group of neoplasms. In 14 patients with UPA multiple tumors, obtained at autopsy, were analyzed by molecular allelotyping and immunohistochemistry. In addition, tumors of 4 patients could be analyzed by comparative genomic hybridization (CGH). Similar genetic and phenotypic profiles were used as indicator for a clonal relationship, whereas different profiles implicate independent tumors. The molecular data indicated that the multiple lesions in the 14 UPA patients, including the primary tumors, are clonally related. In agreement with the theory of tumor progression, some metastatic lesions showed additional genetic alterations besides the characteristics that were shared with the primary tumor. Furthermore, 8 UPA patients had tumors with a high frequency of allelic loss and/or imbalance (FALI; 43-71%), while 6 patients demonstrated a lower FALI (14-29%), suggesting the occurrence of chromosomal instability in the former group. Our data provide molecular evidence for a clonal relationship between multiple metastases and the primary tumor within individual UPA patients, independent of the anatomical origin of the cancer. This finding is in agreement with the suggestion that tumor progression is rapid in UPA patients, limiting the chance of clonal divergence. The identification of 2 groups of UPAs with either a high or low FALI indicates that chromosomal instability is not the only driving force behind early tumor cell dissemination. Thus, other molecular mechanisms must underlie the common biology of these tumors.

Antineoplastic chemotherapy in Jehovah's Witness patients with acute myelogenous leukemia refusing blood products - a matched pair analysis.

BACKGROUND: Acute myelogenous leukemia (AML) may be cured in a substantial number of patients using intensive chemotherapeutic regimens leading to temporary severe myelosuppression. Patients belonging to the denomination of Jehovah's Witnesses (JW), however, are bound by their religious convictions not to accept blood products and are therefore at higher risk for life-threatening events. Reports how to handle this challenge are mainly anecdotal. MATERIAL AND METHODS: We here report in much more detail about our experience with nine patients belonging to the denomination of JW who were treated for AML in our department from 1998 to 2007 and who explicitly wished to receive chemotherapy without blood transfusions. RESULTS: Reduced dose induction chemotherapy administered by several treatment cycles to prevent sustained myelosuppression still led to complete remissions in three out of nine of JW patients but was associated with a high rate of relapse. No durable remission was achieved. The overall hazard ratio for death was 12.1 compared to a matched control group treated with full transfusion support. The predominant cause of non-AML mortality was severe anemia (four out of five early deaths) and uncontrollable bleeding (n = 1). CONCLUSION: Reduced dose chemotherapy without transfusion support in JW suffering from AML is associated with a lower rate of remission, high mortality by severe anemia and very low chances for long-term remissions. Less hematotoxic treatment options including hypomethylating agents or molecular targeted therapies with intensive consolidation after improvement of bone marrow function are promising for these patients but need further investigation.

Intraperitoneal application of rituximab in refractory mantle cell lymphoma with massive ascites resulting in local and systemic response.

In the past decade, rituximab in combination with polychemotherapy has become the standard approach in most patients with advanced CD20-positive B-cell lymphoma. In mantle cell lymphoma (MCL), follicular lymphoma and diffuse large B-cell lymphoma, rituximab has been used as monotherapy and in combination with various chemotherapy regimens in different treatment situations. Routinely, rituximab is given intravenously, but other routes of administration have also been described. Here, we report a 64-yr-old woman who was previously treated with three different chemotherapy regimens for stage IV MCL. No sustained clinical and radiological response could be achieved. The patient's general status declined and she developed massive ascites as the dominant clinical problem. Local, intraperitoneal administration of rituximab was initiated as an experimental treatment approach. After 11 doses of rituximab, the general status of the patient improved significantly, ascites resolved completely and computed tomography (CT) scans demonstrated a partial remission of intra-abdominal lymph nodes and splenomegaly. Furthermore, we observed a regression of mediastinal lymph nodes, pleural effusion and centrocytes in peripheral blood as well as improvement of anaemia. The response to the experimental treatment has maintained for more than 6 months. In summary, we observed a sustained local and systemic response to intraperitoneal administration of rituximab in a patient with heavily pretreated advanced MCL.

Inactivation of the tissue inhibitor of metalloproteinases-2 gene by promoter hypermethylation in lymphoid malignancies.

The tissue inhibitor of metalloproteinases-2 (TIMP-2) is known to antagonize matrix metalloproteinase activity and to suppress tumor growth, angiogenesis, invasion and metastasis. We analysed the methylation status of the CpG island in the TIMP-2 promoter region by methylation-specific polymerase chain reaction (MSP) in hematopoietic cell lines. TIMP-2 promoter hypermethylation in the lymphoma cell line Raji and the leukemia cell line KG1a was associated with transcriptional repression. Treatment with the demethylating agent 5-aza-2'-deoxycytidine resulted in TIMP-2 upregulation in both cell lines. TIMP-2 was expressed in the cell lines HL60, U266 and XG1, which carry an unmethylated promoter region. MSP analysis of primary patient samples revealed aberrant methylation of TIMP-2 in 33/90 (36.7%) cases of non-Hodgkin's lymphoma (NHL), but not in normal peripheral blood lymphocytes as well as in nonmalignant bone marrow and lymph nodes. The frequency of TIMP-2 methylation was slightly higher in aggressive NHL subtypes compared to those with an indolent subtype (38.6 versus 33.3%). In contrast, TIMP-2 was not hypermethylated in any of the 40 cases of acute myelogenous leukemia examined. We conclude that promoter hypermethylation of TIMP-2 is a novel epigenetic event in the pathogenesis of lymphoid malignancies and may contribute to a more aggressive NHL phenotype.

Recombinant CD64-specific single chain immunotoxin exhibits specific cytotoxicity against acute myeloid leukemia cells.

CD64, the high affinity receptor for IgG (FcgammaRI) is expressed on acute myeloid leukemia blast cells and has recently been described as a specific target for immunotherapy. To generate a recombinant immunotoxin, the anti-CD64 single chain fragment (scFv) m22 was cloned into the bacterial expression vector pBM1.1 and fused to a deletion mutant of Pseudomonas exotoxin A (ETA'). Genetically modified Escherichia coli BL21 Star (DE3) were grown under osmotic stress conditions in the presence of compatible solutes. After isopropyl beta-D-thiogalactoside induction, the 70-kDa His(10)-tagged m22(scFv)-ETA' was directed into the periplasmic space and purified by a combination of metal-ion affinity and molecular size-chromatography. The characteristics of the recombinant protein were assessed by ELISA, flow cytometry, and toxicity assays, using CD64-positive AML cells. Binding specificity of m22(scFv)-ETA' was verified by competition with the parental anti-CD64 monoclonal antibody m22. The recombinant immunotoxin showed significant toxicity toward the CD64-positive cell lines HL-60 and U937 reaching 50% inhibition of cell proliferation at a concentration (IC(50)) of 11.6 ng/ml against HL-60 cells and 12.9 ng/ml against U937 cells. Approximately 41% of primary leukemia cells from a patient with CD64-positive AML were driven into early apoptosis by m22(scFv)-ETA' as measured by flow cytometric analysis. This is the first article documenting the specific cytotoxicity of a novel recombinant immunotoxin with major implications for immunotherapy of CD64-positive diseases.

Clinical implications of aberrant DNA methylation patterns in acute myelogenous leukemia.

Hypermethylation of CpG islands near gene promoter regions is associated with transcriptional inactivation and represents an important mechanism of gene silencing in carcinogenesis. Such epigenetic phenomena can act alongside DNA mutations and deletions to disrupt tumor-suppressor gene function. The methylation status of the promoter-associated CpG islands from 11 well-characterized cancer-related genes was analyzed by methylation-specific polymerase chain reaction in 60 adult patients with acute myelogenous leukemia (AML) at diagnosis. The frequency of aberrant methylation among the patient samples was 45.0% (27/60) for suppressor of cytokine signaling-1, 31.7% (19/60) for p15, 20.0% (12/60) for retinoic acid receptor beta2, 13.3% (8/60) for p73 and E-cadherin, 5.0% (3/60) for O(6)-methylguanine DNA methyltransferase, 3.3% (2/60) for death-associated protein kinase 1 and hMLH1, 1.7% (1/60) for p16, and 0% (0/60) for the tissue inhibitor of matrix metalloproteinases-3 and Ras association domain family 1A. Aberrant DNA methylation was found in AML of all French-American-British subtypes and throughout all cytogenetic risk groups. There appeared to be a trend towards a higher methylation frequency in AML patients with an unfavorable karyotype, but this difference was not statistically significant. Our data indicate that hypermethylation of multiple genes involving fundamental cellular pathways is a common event in AML, which varies greatly in frequency among the genes examined. The accumulation of epigenetic events affecting genes which are involved in regulating cell cycle inhibition, cell adhesion, growth factor signaling, and apoptosis may contribute to the malignant AML phenotype. The growing knowledge of the role of epigenetics in the aberrant silencing of cancer-related genes provides a rationale and molecular basis for targeted therapeutic approaches with demethylating agents in AML.

Synergism between rViscumin and cisplatin is not dependent on ERCC-1 expression.

Interactions between recombinant mistletoe lectin (rViscumin) and anticancer drugs were investigated in vitro. rViscumin enhanced the cytotoxic effects of vincristine, mafosfamide, idarubicin and cisplatin in the human leukemia cell lines K562 and KG1a. In human marrow progenitor cells, rViscumin inhibited colony formation and did not exert any protective activity against cisplatin-induced inhibition of clonogenicity. Quantitative real-time reverse transcription polymerase chain reaction analysis revealed that cisplatin treatment of K562 cells resulted in a 1.9-fold increase in mRNA expression of the nucleotide excision repair gene ERCC-1. This upregulation was not prevented when cells were post-incubated with rViscumin. Our study provides evidence that rViscumin is capable of enhancing cytotoxicity of anticancer agents in vitro. This synergism appears to be independent of transcriptional activity of DNA repair relevant genes.

Interleukin-6 affects melphalan-induced DNA damage and repair in human multiple myeloma cells.

In addition to signaling proliferation, growth factors may contribute to the persistence of hematopoietic tumors upon chemotherapeutical challenge. In multiple myeloma, malignant growth is mediated either by paracrine interleukin-6 (IL-6) elaborated by bone marrow stromal cells or via autocrine loops by malignant myeloma cells themselves. Although melphalan is one of the most active drugs in this tumor entity, the development of drug resistance frequently impedes cure of patients by attenuating melphalan-induced DNA-damage. We analyzed whether IL-6 protects XG-1 cells and plasma cells of a patient suffering from end-stage multiple myeloma (plasma cell leukemia) from melphalan with respect to DNA damage and DNA repair. Investigating the housekeeping gene glucose-6-phosphate dehydrogenase (G6PD) by using a PCR-stop assay, we found that there was more DNA damage in the G6PD gene of IL-6 deprived XG-1 and the patient's plasma cells, respectively, than in those with IL-6 supplementation. After cessation of melphalan exposure and 24 hours post-incubation in melphalan-free medium, DNA repair was observed in the patient's plasma cells but not in XG-1 cells. There was more DNA repair with IL-6 addition than without IL-6 addition. Similarly, the apoptotic cell fractions, as measured by flow cytometry, were lower if IL-6 was added to the medium. These results indicate that IL-6 may contribute to drug resistance in multiple myeloma.

[HbH disease--a rare differential diagnosis in a patient with anemia and abdominal pain]. / HbH-Krankheit--eine seltene Differentialdiagnose bei Anämie und Oberbauchschmerzen.

CASE REPORT: A 45-year-old Vietnamese male was admitted to hospital with severe hypochromic anemia and acute abdominal pain. The peripheral blood smear showed extreme anisocytosis and poikilocytosis as well as teardrops and target cells. Hemoglobin electrophoresis and brilliant cresyl blue staining revealed hemoglobin H (HbH) disease with an infection-associated hemolytic crisis. CONCLUSION: In the diagnostic workup of hemolytic and hypochromic anemia, HbH disease as a special type of alpha-thalassemia should be considered early. In patients from Eastern Asia with a mean corpuscular hemoglobin (MCH) < 25 pg, hemoglobin analysis should be performed in order to avoid unnecessary diagnostic procedures. The prognosis of HbH disease is generally favorable, and symptomatic treatment is only recommended during hemolytic crises in association with, e.g., infections or pregnancy.

Solitary plasmacytoma presenting as an intramedullary mass of the cervical cord.

Extramedullary plasmacytomas originating in the spinal cord are extremely rare lesions. We report a 52-year-old man presenting with progressive tetraparesis. Spinal magnetic resonance imaging (MRI) showed an ill-defined contrast-enhancing intramedullary mass at the C5-6 level. After a biopsy was taken, pathology including immunohistochemistry revealed plasmacytoma. Complete workup for multiple myeloma was negative. The patient was treated with intravenous chemotherapy followed by radiotherapy and corticosteroids without any improvement of the clinical symptoms or decreased tumor size on the follow-up MRIs. The most common spinal manifestation of plasmocytoma is involvement of the vertebral bodies leading to extradural compression of the cord. Only rarely plasmocytoma manifests itself as a primarily dural lesion leading to an intradural cord compression. This report describes the extremely rare manifestation of plasmocytoma of the spinal cord itself that therefore should be considered in the differential diagnosis of spinal cord mass lesions.

CpG island methylation patterns in chronic lymphocytic leukemia.

Chronic lymphocytic leukemia (CLL) is the most common leukemia in Western countries. In CLL, a large number of genes affecting cancer-related pathways may be dysregulated by epigenetic silencing. We analysed by methylation-specific polymerase chain reaction the CpG island methylation status of 15 well-characterised cancer-related genes in 32 patients with CLL. Aberrant methylation in the sample of patients with CLL was shown for secreted frizzled-related protein 1 (68.8%), secreted frizzled-related protein 2 (65.6%), death-associated protein kinase 1 (50.0%), E-cadherin (21.9%), secreted frizzled-related protein 4 (15.6%), p15 (9.4%), p16 (6.3%), retinoic acid receptor beta2 (3.1%), secreted frizzled-related protein 5 (3.1%) and tissue inhibitor of matrix metalloproteinases 3 (3.1%). For human Mut-L homolog 1, O(6)-methylguanine DNA methyltransferase, p73, suppressor of cytokine signalling 1 and tissue inhibitor of matrix metalloproteinases 2 no hypermethylation was detected. Hypermethylation of at least one gene was observed in 87.5% of the samples. Our results show that aberrant CpG island methylation affecting cancer-related pathways such as Wnt signalling, regulation of apoptosis, cell cycle control and tissue invasion is a common phenomenon in CLL. Epigenetic disturbances may be involved in the pathogenesis of CLL and thus may provide a molecular rationale for therapeutic approaches.

Aberrant DNA methylation of the transcription factor C/EBPalpha in acute myelogenous leukemia.

We investigated the methylation status of the CCAAT/enhancer binding protein alpha (C/EBPalpha) promoter region near the transcription start site in acute myelogenous leukemia (AML). In hematopoietic tumor cell lines, CpG island hypermethylation of the proximal C/EBPalpha promoter region was associated with transcriptional silencing, and treatment with the demethylating agent 5-aza-2'-deoxycytidine resulted in C/EBPalpha reexpression and promoter demethylation. Aberrant methylation of the C/EBPalpha promoter region occurred in 10/80 diagnostic AML samples, and there was an inverse correlation between aberrant methylation of C/EBPalpha and the negative cell cycle regulator p15. Our results provide further evidence for epigenetic dysregulation of C/EBPalpha in AML.

Impact of angiotensin I converting enzyme inhibitors and angiotensin II type 1 receptor blockers on survival in patients with advanced non-small-cell lung cancer undergoing first-line platinum-based chemotherapy.

PURPOSE: The renin-angiotensin system plays a crucial role in maintaining vascular homeostasis. Stimulation of angiotensin II type 1 receptors (AT1R) acts proangiogenically by increasing levels of vascular endothelial growth factor (VEGF). Consequently, cell culture experiments and animal studies have shown antiproliferative effects of AT1R blockers (ARB) and angiotensin I converting enzyme inhibitors (ACEI) in several malignancies. Until now, very limited clinical data for this antiangiogenic effect exists for combinations with antineoplastic chemotherapy. METHODS: A total of 287 patients with advanced non-small-cell lung cancer undergoing first-line platinum-based chemotherapy were retrospectively analysed regarding long-term medication with ACEI and ARB as well as histological type, stage, performance status, gender, age, dose-intensity of chemotherapy and survival. RESULTS: Patients receiving either ACEI or ARB had a 3.1 months longer median survival than non-recipients (11.7 vs. 8.6 months, HR 0.56, P = 0.03). This survival advantage could not be attributed to other established risk-factors or dose intensity of chemotherapy. CONCLUSIONS: Addition of ACEI or ARB to platinum-based first-line chemotherapy may contribute to prolonged survival in patients with advanced lung cancer.

Targeted restoration of down-regulated DAPK2 tumor suppressor activity induces apoptosis in Hodgkin lymphoma cells.

Death-associated protein kinase 2 (DAPK2) is a calcium/calmodulin-regulated proapoptotic serine/threonine kinase that acts as a tumor suppressor. Here we show that DAPK2 is down-regulated in Hodgkin lymphoma-derived tumor cell lines and that promoter-region hypermethylation is one mechanism for DAPK2 inactivation. To determine whether selective reconstitution of DAPK2 catalytic activity in these cells could induce apoptosis, we created a fusion protein comprising a human CD30 ligand conjugated to a human DAPK2 calmodulin-deletion mutant. Thus, recombinant immunokinase DAPK2'-CD30L has a constitutive kinase activity with enhanced proapoptotic function. We show that this immunokinase fusion protein inhibits cell proliferation and induces apoptotic cell death specifically in CD30/DAPK2-negative tumor cell lines. This proof-of-concept study provides the first demonstration of therapeutic strategies based on the restoration of a defective, tumor-suppressing kinase activity by a novel class of recombinant immunotherapeutics.

Rituximab-associated changes in platelet count in patients with non-Hodgkin lymphoma.

Rituximab is a monoclonal antibody specific for the CD20 antigen. Clinical factors associated with thrombocytopenia after administration of rituximab have only been reported as case reports. We have analysed retrospectively the change of platelet counts following the administration of rituximab in 253 patients with non-Hodgkin lymphoma (NHL). Correlations with clinical and laboratory parameters were assessed. A mean overall decrease in platelets was observed after rituximab infusion. A downward trend in platelet count of more than 30% was observed in 7.2% of the patients. The decline was observed when rituximab was given as a single agent or in combination with chemotherapy. The risk factors to develop a decline in platelets after infusion of rituximab were pre-existent thrombocytopenia, advanced lymphoma stage, bone marrow infiltration, splenomegaly, leukemic presentation, and Burkitt lymphoma histology. In conclusion, a decline in platelet count after administration of rituximab was observed in patients with NHL, mainly those with pre-existing thrombocytopenia.