Rapid and sensitive detection of CpG-methylation using methyl-binding (MB)-PCR

Methylation of CpG islands is associated with transcriptional repression and, in cancer, leads to the abnormal silencing of tumor suppressor genes. We have developed a novel technique for detecting CpG-methylated DNA termed methyl-binding (MB)-PCR. This technique utilizes a recombinant protein with high affinity for CpG-methylated DNA that is coated onto the walls of a PCR vessel and selectively captures methylated DNA fragments from a mixture of genomic DNA. The retention and, hence, the degree of methylation of a specific DNA fragment (e.g. a CpG island promoter of a specific gene) is detected in the same tube by gene-specific PCR. MB-PCR does not require bisulfite treatment or methylation-sensitive restriction and provides a quick, simple and extremely sensitive technique allowing the detection of methylated DNA, in particular in tumor tissue or tumor cells from limited samples. Using this novel approach, we determined the methylation status of several established and candidate tumor suppressor genes and identified the ICSBP gene, encoding the myeloid and B-cell-specific transcription factor interferon consensus sequence-binding protein, as a target for aberrant hypermethylation in acute myeloid leukemia

The Impact of a Tumor Diagnosis on Patients' Attitudes toward Advance Directives

Background: Although advance care planning and the completion of advance directives (ADs) are important tools to avoid unwanted aggressive care once patients have lost their decision-making capacity, only a minority of cancer patients are admitted with completed ADs, and little is known about patients' wishes regarding AD consultations. Methods: For 1 year, every new patient admitted to the hematology/oncology outpatient clinic of the University Hospital Regensburg received a self-administered questionnaire comprising a self-evaluation of AD knowledge and questions about preferences regarding consultation partners and the time of consultation. Disease-related data were collected from medical records. Statistics were calculated with SPSS. Results: Of the 500 questionnaires handed out, 394 (75%) were evaluable and analyzed. Twenty-eight percent of the participants had completed an AD (living will or health care proxy). Ninety-two percent of the participants without ADs had never received a consultation offer from any professional involved. Only 20% perceived a clear relation between cancer and AD consultations. More than 50% of the participants without ADs were in favor of consultations ‘now' or ‘in a few weeks', while more than 40% objected to AD consultations. Conclusions: Oncology patients have a large unmet demand for AD consultations. However, a relevant percentage of these patients object to AD consultations. Structured and early AD consultation offers should be made, and early discussions about indications for aggressive treatment should take place

Prophylactic Application of Nebulized Liposomal Amphotericin B in Hematologic Patients with Neutropenia

Background: Pulmonary invasive fungal infections (IFI) are well-recognized complications with high morbiditiy and mortality in patients with hematologic malignancies. Patients and Methods: Aerosolized liposomal amphotericin B (lipAmB) was evaluated as an antifungal prophylaxis in patients with an expected neutropenia of more than 10 days due to intensive chemotherapy or stem cell transplantation, in a prospective phase II trial. Results: 98 treatment episodes were included in the study and compared to 105 historical control patients. Inhalation was performed between 0 and 103 days. No severe side effects of therapy occurred. 40 patients considered inhalations as unpleasant, 2 as very unpleasant, mostly due to bad taste or cough. Few cases of definite or probable IFI were recorded, whereas a large number of patients were treated with systemic antifungal therapy for pneumonia or fever of unknown origin without a significant difference between study patients and controls. In a predefined subgroup analysis of 48 patients with newly diagnosed acute myeloid leukemia (AML), significantly more patients survived for 1 year in the AmB prophylaxis than in the control group (80% vs. 54%, p < 0.01). Conclusions: Inhalations of lipAmB are feasible and safe. Results in the subgroup of patients with AML together with data from other trials suggest further evaluation of effectiveness

C-reactive Protein in Patients with Metastatic Clear Cell Renal Carcinoma: An Important Biomarker for Tumor-associated Inflammation

Two consecutive multi-center phase II trials were designed to prove the hypothesis, whether therapeutic modeling of tumor-associated inflammatory processes could result in improved tumor response

Papillary thyroid cancer associated with syndrome of inappropriate antidiuresis: a case report

<p>Abstract</p> <p>Introduction</p> <p>The syndrome of inappropriate antidiuresis is the most common cause of euvolemic hypo-osmolality. This syndrome is associated with a wide variety of diseases. However, its most frequent causes are related to malignancies, especially lung cancer. In this case report, we describe an unknown association of the syndrome of inappropriate antidiuresis with papillary thyroid cancer.</p> <p>Case presentation</p> <p>We present the case of a 71-year-old Caucasian, German woman with marked hyponatremia and neurological symptoms. After a detailed clinical investigation, the common causes of syndrome of inappropriate antidiuresis and other malignancies were ruled out. A thyroid nodule was detected by ultrasound and magnetic resonance imaging. Although fine needle aspiration cytology showed negative results, our patient underwent surgery. Papillary thyroid cancer was later diagnosed. After total thyroidectomy, a complete remission of the clinical symptoms occurred and our patient subsequently had iodine-131 radioactive therapy. Hyponatremia was no longer observed during the follow-up investigations.</p> <p>Conclusion</p> <p>This is the first reported case of paraneoplastic syndrome of inappropriate antidiuresis caused by papillary thyroid carcinoma. Since its symptoms occurred before the development of local symptoms, total thyroidectomy may provide a timely and efficient treatment for the underlying malignancy.</p

Induction of human monocyte to macrophage maturation in vitro by 1,25-dihydroxyvitamin D3

Cells of the mononuclear phagocyte system arise from circulating blood monocytes (MO) that undergo further maturation on leaving the vasculature and migration into the various tissues and body cavities. This terminal differentiation step is also observed in vitro when blood MO are cultured in the presence of serum. Yet, the inducing signals present in serum are not defined. We have established primary cultures from elutriation-purified blood MO and found that the active metabolite of vitamin D3 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) could induce maturation of MO to macrophages (MAC) in the absence of any serum proteins. Cells were cultured for 7 days with AB-group serum or 1,25(OH)2D3, respectively, and MO maturation analyzed by morphology, functional activity, and the expression of lineage-restricted maturation-associated antigens (MAX.1, MAX.3). At an optimal concentration of 10(-8) mol/L, 1,25(OH)2D3 promoted the development of fully differentiated MAC whose phenotype and functional competence in terms of cytokine release (tumor necrosis factor alpha, interleukin-6, fibronectin, and lysozyme) was comparable with MAC grown in serum. In conclusion, our data may add to the immunoregulatory potential of 1,25(OH)2D3, which may play an essential role in the ontogeny of the mononuclear phagocyte system

Macrophage colony-stimulating factor is required for human monocyte survival and acts as a cofactor for their terminal differentiation to macrophages in vitro

Functional competence as well as phenotype heterogeneity of macrophages depend on the completion of their maturation pathway. Differentiation of committed myeloid progenitor cells is induced by colony-stimulating factors (CSF), but no consistent data exist on which factor(s) induce the terminal maturation from the circulating blood monocyte to the mature macrophage. In vitro, monocyte to macrophage transformation occurs in the presence of serum and can be followed by the expression of the maturation-associated antigens gp65-MAX.1, gp68-MAX.3, and CD51. We describe that the differentiation-inducing activity in serum cannot be replaced by any of the known and available purified recombinant cytokines. In the absence of serum monocytes die in suspension cultures while surviving as non-differentiating cells when cultured adherent to plastic. In serum-free suspension cultures survival can be significantly improved by the addition of recombinant human macrophage (rhM)-CSF whereas other cytokines do not. At any stage of serum-free adherent culture, monocyte to macrophage differentiation can be induced rapidly by the addition of serum, whereas cytokines (rhM-CSF, recombinant human granulocyte macrophage [rhGM]-CSF, recombinant human granulocyte [rhG]-CSF, recombinant human interleukin [rhIL]-1, rhIL-3, rhIL-4, rhIL-6, tumor necrosis factor [TNF]-alpha, interferon [IFN]-alpha, IFN-gamma) alone or in combination are not effective. Serum-induced maturation, however, was suppressed in the presence of neutralizing anti-M-CSF antibodies. In addition to phenotype analysis, the secretory repertoire of rhM-CSF cultured monocytes was analyzed in comparison to serum cultured monocytes which further characterized them to be immature cells, i.e., low release of maturation-associated products such as alpha-2-macroglobulin, neopterin, fibronectin, and TNF-alpha, but high IL-6 secretion, an attribute of blood monocytes. We conclude that for monocyte survival in vitro the presence of endogenous M-CSF and possibly other autocrine factors elicited by cell adherence are required for the induction of macrophage maturation; however, yet undefined additional factor(s) are necessary. They are present in serum and may act in conjunction with M-CSF but are distinct from all known cytokines. Our in vitro system may be useful in the screening and discovery of these serum factor(s)